Reagent container for an automated analyzer

ABSTRACT

An analyzer for performing automated assay testing. The analyzer includes a storage and conveyor system for conveying cuvettes to an incubation or processing conveyor, a storage and selection system for test sample containers, a storage and selection system for reagent containers, sample and reagent aspirating and dispensing probes, a separation system for separating bound from unbound tracer or labeled reagent, a detection system and date collection/processing system. All of the sub-units of the machine are controlled by a central processing unit to coordinate the activity of all of the subunits of the analyzer. The analyzer is specificallysuited for performing heterogeneous binding assay protocols, particularly immunoassays.

This Appln. is a cont. of Ser. No. 08/457,702 filed Jun 1, 1995, whichis a Div. of Ser. No. 08/222,559 filed Apr. 1, 1994, abnd., which is acont. of Ser. No. 07/665,196 filed Mar. 4, 1991, abnd.

BACKGROUND OF THE INVENTION

The present invention is generally directed to an automated analyzer forconducting binding assays of various liquids, particular biologicalfluids for substances contained therein.

The present invention is particularly directed to a machine forperforming automated immunoassay testing, in particular heterogeneousimmunoassays in which paramagnetic particles are the solid phase reagentand the labeled reagent (tracer reagent) includes a chemiluminescentlabel. The system can accommodate both competitive and sandwich-typeassay configurations. A chemiluminescent flash is initiated and itsintensity measured as an indication of the presence or absence of ananalyte in the test fluid which is being assayed. The analyzer can beselectively run in batch-mode or random access sequence.

Over the last several years, automated instrumentation has beendeveloped for routine testing in the clinical laboratory. Limitedautomation has been applied to the area of immunoassay testing. Althoughsome instruments have been developed for limited immunoassay testing,many of the procedures are still performed manually. Test results arevery often delayed because of the time factor and labor intensity formany of the manual steps, and long incubation or reaction times. Thesedelays can be critical in many clinical situations. In addition, themanual procedures cause variations in test results and are quite costly.The causes of such variations include nonuniform testing protocols,technician experience skills and the precision of theapparatus/analyzer. These and other difficulties experienced with theprior art analyzer and manual testing systems have been obviated by thepresent invention.

It is, therefore, a principal object of the invention to provide anautomated analyzer for diagnostic immunoassay testing which isparticularly applicable to heterogeneous immunoassay testing.

Another object of this invention is the provision of an analyzer whichhas a high degree of versatility, capable of performing a wide range ofbinding assay protocols for a wide range of clinical and non-clinicalanalytes.

A further object of the present invention is the provision of anautomatic analyzer which is capable of handling a plurality of testprotocols simultaneously, continuously and sequentially.

It is another object of the present invention to provide an automatedanalyzer which is capable of high sample throughput.

A still further object of the invention is the provision of an automatedanalyzer which greatly reduces the amount of time per assay or sampletest.

It is a further object of the invention to provide an automated analyzerwhich provides consistent and reliable assay readings.

It is a further object of the invention to provide an automated analyzerwhich is self-contained and requires a minimal amount of space forcomplete sample processing.

A further object of the invention is to provide a constant luminescentlight source for automatic monitoring of the luminometer calibration ofan assay apparatus.

It is still a her object of the invention to provide an automatedanalyzer which can be selectively run in a bath-mode or random accesssequence.

With these and other objects in view, as will be apparent to thoseskilled in the art, the invention resides in the combination of partsset forth in the specification and covered by the claims appendedhereto.

SUMMARY OF THE INVENTION

In general, the automated analyzer of the present invention is aself-contained instrument which is adapted to be located on a suitablelaboratory bench. It requires no external connections other than astandard power line and operates accurately within an ambienttemperature range of 18° to 30° C. The functional units of the analyzerinclude a process track, a sample handling or transport system, areagent handling or transport system, a separation and washing system, adetection system (luminometer) and data collection/processing system.The reagents and test samples are reacted in discreet, disposablecuvettes. The cuvettes are automatically and sequentially dispensed froma cuvette loader onto a linear process tract which moves each cuvetteone cuvette space every twenty seconds. The temperature of the testreaction is controlled by a thermal system which preheats the cuvettesand reagent and maintains an environmental temperature of 37° C., plusor minus one degree, throughout incubation. Test samples are dispensedinto the cuvettes by an aspirating and dispensing probe and reagents areadded at software-controlled intervals by means of three aspirating anddispensing reagent probes. The analyzer is particularly adapted forperforming heterogeneous specific binding assays. The analyzer can beselectively run in batch-mode or random access sequence.

BRIEF DESCRIPTION OF THE DRAWINGS

The character of the invention, however, may be best understood byreference to one of its structural forms, as illustrated by theaccompanying drawings, in which:

FIG. 1 is a front perspective view of the analyzer of the presentinvention;

FIG. 2 is a diagrammatic plan view showing the general organization ofthe subunits of the analyzer;

FIG. 3 is a diagrammatic plan view of a sequential series of cuvetteswhich are disposed on the pre-heater section and event conveyor;

FIG. 4 is a front elevational view of a cuvette which is used with theautomated analyzer of the present invention for holding sample andreagent;

FIG. 5 is a top plan view of the cuvette;

FIG. 6 is a bottom plan view of the cuvette;

FIG. 7 is a side elevations view of the cuvette;

FIG. 8 is a perspective view of the cuvette;

FIG. 9 is a side elevational view of a container for holding reagent,specifically labeled reagent (tracer reagent);

FIG. 10 is a top plan view of the container;

FIG. 11 is a bottom plan view of the container;

FIG. 12 is a perspective view of the container;

FIG. 13 is a vertical cross-sectional view of the container taken alongthe line 13--13 and looking in the direction of the arrows;

FIG. 14 is a bottom plan view of a cover for a container including thecontainer which is shown in FIG. 9;

FIG. 15 is a vertical cross-sectional view of the cover taken along theline 15--15 and looking in the direction of the arrows;

FIG. 16 is a side elevational view of a reagent container, specificallyfor solid phase reagent;

FIG. 17 is a top plan view of the solid phase reagent container;

FIG. 18 is a bottom plan view of the reagent container;

FIG. 19 is a vertical cross-sectional view of the reagent container,taken along the line 19--19 of FIG. 17 and looking in the direction ofthe arrows;

FIG. 20 is a perspective view of the reagent container with portionsbroken away;

FIGS. 21A and 21B, when viewed together, is a front elevational view ofthe analyzer of the present invention, the sheet being joined along theline 21A;

FIG. 22 is a top plan view of the analyzer, with portions broken away;

FIG. 23 is an end view of the analyzer;

FIG. 24 is an exploded perspective view of a system for feeding cuvettesfrom a storage hopper;

FIG. 25 is a perspective view of a cuvette storage hopper;

FIG. 26 is an exploded perspective view of the cuvette feed system andhopper;

FIG. 27 is a front elevational view of the cuvette feed system;

FIG. 28 is a rear elevational view of the cuvette feed system;

FIG. 29 is a right side elevational view of the cuvette feed system,with portions broken away;

FIG. 30 is a plan view of the hopper and feed system;

FIG. 31 is a fragmentary view of a feed chute which forms part of thecuvette feed system, with portions broken away;

FIGS. 32A, 32B and 32C, when taken together, form a front view of aconveyor system for feeding cuvettes from the hopper feed system throughthe vent areas of the machine, the sheets being joined along the lines32A and 32B;

FIGS. 33A, 33B and 33C, when viewed together, form a top plan view ofthe cuvette conveyor system the sheets being joined along the lines 33Aand 33B;

FIG. 34 is a vertical cross-sectional view showing magnetic means forattracting paramagnetic particles from the test sample and reagentmixture in a cuvette taken along the line 34A--34A of FIG. 33C andlooking in the direction of the arrows;

FIG. 35 is a vertical cross-sectional view showing another aspect of themagnetic means for attracting the paramagnetic particles from the testsample and reagent mixture within a cuvette taken along the line35A--35A of FIG. 33C and looking in the direction of the arrows;

FIG. 36 is a front elevational view of a sample transport system;

FIG. 37 is a top plan view of the sample transport system;

FIG. 38 is a vertical cross-sectional view of the sample transportsystem taken along the line 38A--38A of FIG. 37;

FIG. 39 is an exploded perspective view of some of the elements of thesample transport system;

FIG. 40 is an exploded perspective view of one of the drive mechanismsfor the sample transport system;

FIG. 41 is an exploded diagrammatic elevational view of the sampletransport system;

FIG. 42 is a perspective view of one of the drive elements of the sampletransport system;

FIG. 43 is a top plan view of a reagent transport system;

FIG. 44 is a front elevational view of a reagent transport system;

FIG. 45 is a vertical cross-sectional view of the reagent transportsystem;

FIG. 46 is an exploded perspective view of some of the elements of thereagent transport system;

FIG. 47 is an exploded perspective view of additional elements of thereagent transport system;

FIG. 48 is an exploded perspective view of one of the drive elements forthe reagent transport system;

FIG. 49 is a diagrammatic elevational view of the reagent transportsystem;

FIG. 50 is a front elevational view of a sample probe transport system;

FIG. 51 is a diagrammatic right side elevational view of the sampleprobe transport system;

FIG. 52 is a right side elevational view of the sample probe transportsystem;

FIG. 53 is a plan view of the sample probe transport system;

FIG. 54 is an exploded perspective view of some of the elements of thesample probe transport system;

FIG. 55 is an exploded perspective view of the horizontal drivecomponents of the sample probe transport system;

FIG. 56 is an exploded perspective view of a sample probe supportingcarriage which forms part of the sample probe transport system;

FIG. 57 is an exploded elevational view of one of the drive componentsfor the sample probe transport system;

FIG. 58 is an exploded perspective view of one of the horizontal drivecomponents for the sample probe transport system;

FIG. 59 is an exploded perspective view of one of the vertical drivecomponents for the sample probe transport system;

FIG. 60 is a top plan view of a reagent probe transport system;

FIG. 61 is a right side elevational view of the reagent probe transportsystem;

FIG. 62 is a front elevational view of the reagent probe transportsystem;

FIG. 63 is an exploded perspective view of some of the elements of thereagent probe transport system;

FIG. 64 is an exploded perspective view of the components of the lefthand reagent probe;

FIG. 65 is an exploded perspective view of the central reagent probecomponents;

FIG. 66 is an exploded perspective view of the right reagent probecomponents;

FIG. 67 is an exploded perspective view of one of the horizontal driveelements of the reagent probe transport system;

FIG. 68 is an exploded perspective view of one of the drive componentsfor moving the left probe vertically;

FIG. 69 is an exploded perspective view of the probe supporting elementsfor the central probe of the reagent probe transport system;

FIG. 70 is an elevational view of a post which forms part of themechanism for rotating the left probe about a vertical axis;

FIG. 71 is an exploded perspective view of the probe supporting elementsfor the right probe of the reagent probe transport system;

FIG. 72 is an exploded perspective view of the probe supporting elementsfor the left probe of the reagent probe transport system;

FIG. 73 is an exploded perspective view of the syringe bank for thesample and reagent probes;

FIG. 74 is a cross-sectional view of a heating system for a tube whichextends from one of the reagent probes to its corresponding syringe;

FIG. 75 is an exploded perspective view of an event conveyor system andall of the wash stations for the sample and reagent probes;

FIG. 76 is a perspective view of the right hand end of the analyzerwhich illustrates the aspirate resuspend area of the event track and theluminometer;

FIG. 77 is an exploded perspective view of the aspirate resuspendcomponents;

FIG. 78 is a cross-sectional view of one of the aspirating probes;

FIG. 79 is a vertical cross-sectional view of a cuvette wash apparatuswhich forms part of the aspirate resuspend section of the event conveyortaken along the line 79A--79A of FIG. 33C;

FIG. 80 is a vertical cross-sectional view of the acid resuspendmechanism taken along the line 80A--80A of FIG. 33C;

FIG. 81 is a right hand elevational view of a luminometer and elevatormechanism which conveys cuvettes to the luminometer at the end of theevent conveyor;

FIG. 82 is a top plan view of the luminometer;

FIG. 83 is a vertical cross-sectional view of the luminometer andcuvette elevator;

FIG. 84 is an exploded perspective view of some of the elements of theluminometer;

FIG. 85 is a perspective view of the luminometer;

FIG. 86 is a diagrammatic plan view showing the path of the cuvetteswithin the luminometer;

FIGS. 87A-B is a schematic diagram of a preferred embodiment of areference LED module;

FIG. 88 is a block diagram of the module;

FIG. 89 is a diagram of the preferred timing scheme of an electronicallyadjustable potentiometer in the reference LED module;

FIG. 90 is an exploded perspective view of the valve modules which arelocated at the left side of the analyzer;

FIG. 91 is a perspective view of the left side valve components andperistaltic pumps;

FIG. 92 is an exploded perspective view of the valve components at theright hand side of the analyzer;

FIGS. 93A and 93B is a schematic view of all of the pneumatic andplumbing components for the analyzer;

FIGS. 94-102 are flow diagrams of the coordinated operation of thevarious subunits of the analyzer.

It is noted that the representations shown in the FIGS. may not indicateactual scales or ratios.

GLOSSARY

The following terms as used in this specification and claims are definedas follows:

ACID REAGENT:

0.1N HNO₃ with 0.5% peroxide; added to the magnetic particles after thewash cycle. The peroxide attaches to the acridinium ester at a low pH(pH1). This reaction readies the acridinium ester for light emission.

ACRIDINIUM ESTER (AE):

The chemical "label" responsible for the chemiluminescent flash whenbase reagent is added to the acidified magnetic particle/analyte/AEmixture in the cuvette. See U.S. Pat. Nos. 4,745,181, 4,918,192 and4,946,958, which are incorporated by reference.

ANALYTE:

A substance of unknown concentration present or suspected of beingpresent in a test sample.

ANTIBODY (Ab):

1) a protein produced by the body in response to the presence of aforeign substance; part of the body's resistance to disease 2) proteinsor carbohydrates containing proteins having the ability to combine witha specific antigen.

ANTIGEN (Ag):

1) a substance foreign to the body which when introduced into the bodystimulates the production of antibodies 2) under analysis conditions; aprotein or non-protein compound capable of reacting with a specificantibody.

ASSAY:

a diagnostic or analytical protocol for determining the presence andamount or absence of a substance in a test sample, said away includingimmunoassays of various formats.

BASE REAGENT:

0.25N NaOH, pH 13, and ARQUAD; added to the magnetic particles suspendedin acid when the cuvette is in the luminometer. When injected, the pHshift and accompanying electron excitation causes light emission at aspecific wavelength (a flash). See U.S. Pat. No. 4,927,769 which isincorporated by reference.

BUFFER:

A solution used for pH maintenance; composed of a weak acid (or base)and its salt.

CALIBRATOR:

A protein based solution (often human based) containing knownconcentrations of analytes providing a reference curve for convertingmeasured signal into concentration.

CALIBRATION CURVE:

A pair of calibrators are run as samples and the calibrator data isnormalized against the stored Master Curve data for the tested analyte,compensating for current running conditions and instrument variability.

CHEMILUMINESCENCE:

A chemical reaction in the production of light

COMPETITIVE ASSAY:

An Ab/Ag reaction where the unknown Ag in a sample and a labeled Ag inreagent compete for a limited amount of reagent labeled Ab.

CONTROL:

A protein based product containing specific analytes within apre-determined concentration range; i.e., low, medium, high. Manycontrols are human serum based. Controls are used as a total systemperformance check.

COUNTS:

The basic unit of measurement of PMT signal after processing by the PADelectronics.

COUNT PROFILE:

Counts vs time; information is stored in files in system and can beplotted.

DARK COUNTS:

The electronic noise of the PMT the absence of light.

DILUENT (DIL):

A protein based solution; used to dilute a patient sample when theoriginal result is beyond the curve range.

FLASH:

A short-lived burst of light produced from the immunoassay when the pHis rapidly changed from acidic to basic (with the addition of the basereagent).

HAPTEN:

An incomplete antigen being incapable alone of causing the production ofantibodies but capable of combining with specific antibodies.

IMMUNOASSAY:

A chemical test involving an antibody/antigen reaction to determine thepresence of and/or quantify a specific substance; the substance beingassayed may be the antibody or antigen in the reaction.

LIGHT COUNTS:

The electronic signal of the PMT in the presence of light, includingdark counts.

MASTER CURVE:

A ten point curve generated by Quality Control for each matched set ofSP and Lite reagents, data is published in assay's package insert andprogrammed into instrument by operator; used by instrument as the masterreference curve for converting measured signal into concentration.

NSB:

non-specific binding--All tracer material which is present during themeasurement phase but does not represent specific Ab binding. Tracermaterial may attach indiscriminately to cuvette wall or particles anddoes not wash away, resulting in signal that mimics an Ab/Ag reaction.

PAD:

Electronics that amplify the PMT signal (pulse) and filter it for signalnot generated by photons.

PHOTON:

A unit of light.

PMP:

Para-magnetic particles; used in Solid Phase reagent.

PMT:

Photomultiplier tube--a vacuum (or gas-filled) phototube with a cathode,usually nine dynodes, and an anode. The cathode is capable of emitting astream of electrons when exposed to light. The dynode arrangementprovides successive steps in amplification of the original signal fromthe cathode. The resulting signal produced is directly proportional tothe amount of illumination.

PRE-TREATMENT AGENT (TRX):

A solution mixed and incubated with sample to protect the analyte fromreleasing agent.

RELEASING AGENT (REL):

A solution mixed with sample for the purpose of separating the analytefrom another molecule and rendering it available for immuno-reaction.

RLU:

Relative light units; used on the manual Magic^(R) Lite analyzers. Aunit of light measurement calibrated against a tritium source and uniquefor each instrument.

SANDWICH ASSAY:

An Ab/Ag reaction where unknown Ag reacts with two forms of reagentlabeled Ab; a solid phase or physical carrier reagent and a signalproducing reagent, resulting in a Ab/Ag/Ab "sandwich".

SOLID PHASE REAGENT (SP):

A physical carrier reagent coupled with antigen or antibody (as requiredby assay) in a buffer. See U.S. Pat. Nos. 4,554,088 and 4,672,040.

SYSTEM FLUID (system water, system diluent):

All system syringes are water backed with D.I. water from the on-boardsupply; used to follow sample and reagent dispense to cuvette, wash allprobes, wash magnetic particles in cuvette at aspirate/resuspendposition in track.

TEST SAMPLE:

A specimen for testing; including biological fluids, e.g. serum, urine,cellular products, controls, calibrators, etc., non biological fluids,e.g. chemical compounds, drugs, etc., and any other fluid of interestfor which an assay protocol may be formatted.

TOTAL COUNTS:

1) the area under the flash curve 2) counts per read interval.

TRACER REAGENT (Lite Reagent (LR)):

Antibody or antigen (as required by assay) labeled with acridinium esterin a barbitol buffer (synonym--tracer).

TRITIUM:

A radioactive light source in a sealed scintillation solution; it emitslight and serves as a calibration reference for evaluating luminometerperformance. (Los Alamos Diagnostics product insert; PN 71-002 &61-006).

DESCRIPTION OF THE PREFERRED EMBODIMENT General Organization of MachineSubunits

The analyzer requires onboard supplies of cuvettes, deionized water, andthe acid and base reagents. Sensors monitor volumes of liquid suppliesand indicate necessary refilling before the assay run is initiated.Additional cuvettes may be loaded at any time, even while the instrumentis operating. Waste liquid is collected in an on-board removablereservoir, and used cuvettes are collected in a waste bin, afteraspiration of all liquid waste. The analyzer advises the operator wheneither of these waste collectors are in need of emptying.

Referring first to FIGS. 1, 2 and 3, the automated analyzer of thepresent invention and includes a housing 21 which contains or supports aplurality of subunits for performing the various steps for completion ofa plurality of binding assays on fluid samples, e.g. blood serum. Theanalyzer is specifically adapted to perform heterogeneous immunoassayshaving various formats. The subunits include a cuvette hopper and feedermechanism which is generally indicated by the reference numeral 22, acuvette conveying system 23, a sample probe transport system 24, aplurality of reagent probe transport systems R1, R2 and R3, a sampletransport system which is generally indicated by the reference numeral26, and a reagent transport system which is generally indicated by thereference numeral 27. A detection device 29 is located at the end of andabove the conveyor system 23. The detection device of the preferredembodiment is a luminometer. Other devices, e.g. fluorimeter, isotopeemitter counters, etc. are known in the arts. The uses of such otherdevices is determined by the type of label that is utilized in a testreaction. This system 20 also includes a syringe bank 32, a centralprocessing unit (CPU). not shown, which is operably connected to acathode ray tube (CRT) 36 and keyboard 37. The syringe bank 32 isoperatively connected to the sample probe transport system 24 andreagent probe sport systems R1, R2 and R3.

A wash station for the sample aspirating and dispensing probe is locatedbehind the sample transport system and is generally indicated by thereference numeral 18. Additional wash stations, generally indicated bythe reference numerals 15, 16 and 17, for the reagent aspirating anddispensing probes are located behind the reagent transport system 27,see also FIGS. 21A, 21B and 22.

Referring particularly to FIG. 3, the conveyor system 23 is divided intotwo sections, a cuvette preheater section which is generally indicatedby the reference numeral 38 and a cuvette dispense and incubationsection which is generally indicated by the reference numeral 39. Thecuvettes 40 are stored in a random manner in a hopper 22 and conveyed tothe end of the preheater section 38 in an upright orientation. A plunger19 is fixed to the end of a lead screw 41 which is driven horizontallyby an electric motor 25 along its central longitudinal axis and the axisof the preheater section 38. The plunger 19 is moved from an outerretracted position to an extended position as shown in FIG. 3 to push acuvette which has just been deposited on the preheater section 38 onecuvette space towards the incubation section 39. This advances all ofthe cuvettes 40 along the preheater section 38 so that the furthestcuvette is transferred onto the incubation section 39. The plunger 41 isthen moved back to the retracted position to engage the next cuvettewhich will drop into the starting position. The lead screw 41 does notrotate about its axis. Cuvette sensors, generally indicated by thereference numeral 43, are positioned at the end of the preheat section38 and at the beginning of the incubation section 39 to monitor thepresence of cuvettes at these locations. The cuvettes 40 are conveyedalong the incubation section 39 by conveyor means, described below,which is driven by a motor 42. As each cuvette reaches a sample dispensepoint 44 along the incubation section 39, a probe, described below, fromthe sample probe transport system 24 aspirates a predetermined amount offluid to be analyzed from a container, described below, in the sampletransport system 26 and deposits the sample in the cuvette at the sampledispense point 44. When the cuvette reaches any one of threepredetermined positions 45, 46 or 47 adjacent the reagent transportsystem 27, a pair of reagents from the reagent transport system 27 isadded to the fluid sample in the cuvette to initiate a test reaction forforming a detectable product by one or more of the reagent probes fromthe reagent probe systems R1, R2 or R3. The sequence of reagent additioninto the cuvette is determined by the assay protocol selected for thetest sample. Variation in reagent addition occurs for example when anincubation of test sample and one of the reagents is required. Thereagents comprise a solid phase reagent and a labeled reagent (tracerreagent) which, in the preferred embodiment, is of a luminescentcompound.

The solid phase reagent in the preferred embodiment is paramagneticparticles having a binding substance coupled thereto. Alternate solidphase materials are known in the arts as well as separation techniquesfor isolating the said solid phase materials. The detectable productthat is formed in the preferred embodiment is a complex that includesthe solid phase reagent, analyte that is being assayed and the labeledreagent. The complex will vary depending on the format of the assay.Examples of binding assay formats which generate a detectable productinclude competitive and sandwich type reactions, each of which may beperformed by the analyzer of the present invention. Thereafter, thecuvette passes an aspirate/resuspend area which is generally indicatedby the reference numeral 28, which prepares the mixture for a "flash" orlight emitting reaction in the luminometer 29. Referring particularly toFIG. 3, the aspirate resuspend area 28 of the preferred embodimentincludes a magnetic apparatus 49. An aspirate/wash probe is located atpoint 50. An aspirate probe is located at point 51 and an acidresuspension probe is located at point 52.

When the cuvette reaches the end of the incubation section 39, it islifted vertically by an elevator mechanism at point 53 to theluminometer 29. When the cuvette which contains the acid resuspendeddetectable product has been properly positioned within the luminometer,a base solution is added which results in a chemiluminescent detectionreaction ("flash"). The "flash" effects a photomultiplier tube whichcounts photons from the "flash" and produces an electrical signal. Thesignal is processed by the central processing unit and an appropriatevalue reading is recorded. Deionized water is used for a system backingfluid and for many of the washing steps for typical assay protocols andis are stored in a removable reservoir 30. A second removable reservoir31 is located below the reservoir 30 for accepting all fluid waste.After each assay, the contents of the cuvette are aspirated from thecuvette and discharged into the fluid waste reservoir 31. The emptycuvette is then discarded into a waste receptacle 35. Acid reagent isstored in a reservoir 33 and base reagent is stored in a reservoir 34.An example of an acid reagent which is suitable for use with the presentsystem is: 0.1N. HNO₃, pH 1.0 with 0.5% peroxide. An example of a basereagent which is suitable for use with the present system is 0.25N.,NaOH, pH 13, and ARQUAD. Variations in the concentration of the acid andbase reagents may be required depending on the chemiluminescent label.The chemiluminescent label in the preferred embodiment is an acridiniumester.

Cuvette and Reagent Containers

Referring to FIGS. 4-8, the cuvette which is used as part of theautomated analyzer of the present invention is generally indicated bythe reference numeral 40. Cuvette 40 is generally rectangular incross-section and consists of a bottom wall 55, a pair of opposite broadside walls 56 and a pair of opposite narrow sidewalls 57. The cuvette 40has an interior chamber which is accessed from a top opening 69. A pairof flanges 58 extend outwardly from the broad sidewall 56 at the top ofthe cuvette. A pair of spaced teeth 59 extend outwardly from each broadsidewall 56 just below the flange 58. The flanges 58 and teeth 59 areinstrumental in enabling the cuvette to be conveyed and transportedthrough the various subsystems of the machine 20, as will be describedhereafter. The cuvette can be made of polypropylene or polyethylenewhich have been found to produce a more even light distribution duringthe subsequent flash in the luminometer than other polymers which havebeen tested such as polystyrene. However, polypropylene has been foundto be the preferred material for obtaining reliable results.

Referring to FIGS. 9-13, one of the two types of reagent containerswhich are utilized in the analyzer, is generally indicated by thereference numeral 60. The container 60 is utilized for caring a labeledreagent (tracer reagent) which is specific for certain test protocolsand comprises a main body portion 64 which has an inner chamber 61, athreaded neck portion 65 and a top opening 62 at the upper end of theneck portion 65 which opens into the chamber 61. A skirt 63 extendsoutwardly from a point below the neck 65 and extends downwardly to apoint just below the main body portion 64. The skirt 63 is spaced fromthe main body portion 64 and consists of three flat sides and onerounded side. The skirt 63 enables the container 60 to be securelymounted on the reagent transport means, described below.

FIGS. 14 and 15 illustrate a cover for a container including the reagentcontainer 60 which is generally indicated by the reference numeral 66and includes a top wall 67 which has a plurality of slits 68 which crossat the center of the top wall 67. The cover 66 is made of an elastomericmaterial such as natural or synthetic rubber which enables the cover toengage the top of the neck portion 65 of the container 60. The cover 66reduces evaporation of reagent from the container 60 and the slits 68enable a reagent aspirating and dispensing probe to penetrate the topwall 67 to access the reagent fluid within the container. The slits 68all intersect at the center of the top wall 67 to form a plurality ofpie-shaped flaps which converge at the center of the cover and give waywhen pressure is applied to the center of the cover. The bottom of thecover 66 has an outer annular flange 70.

FIGS. 16-20 illustrate a second reagent container which is used with theanalyzer and which is generally indicated by the reference numeral 75for holding a solid phase reagent. The container 75 has a generallycylindrical main body portion 76 which has an inner chamber 77 whichextends to a top opening 78 above a threaded neck portion 79. An annularskirt 80 extends outwardly from the main body portion 76 at a point justbelow the neck 79 and extends downwardly to a point below the main bodyportion 76, as shown most clearly in FIG. 19. A pair of fins 81 extendinwardly into the chamber 77 from the inner chamber wall as shown mostclearly in FIGS. 17 and 20. The fins 81 are utilized for agitating thesolid phase reagent within the container in a manner described below inconnection with the reagent transport system 27. The top opening 78 isalso sealed by the cover 66 by inverting the cover so that the top wall67 extends below the top opening 78 and inside of the neck portion 79 sothat the flange 70 of the cover rests on top of the neck portion 79.

Cuvette Feed and Orientation Mechanism

Referring to FIGS. 24-31, the cuvette feed and orientation mechanism 22comprises a hopper which is generally indicated by the reference numeral87, a feed conveyor which is generally indicated by the referencenumeral 86, and an orientation chute which is generally indicated by thereference numeral 131. The hopper 87 is preferably made of an opticallyclear plastic material. This makes it easier for the operator todetermine when the level of cuvettes in the hopper is low whereby thehopper requires additional cuvettes. In addition, the elements which arebelow the hopper, see FIG. 30.

Referring particularly to FIGS. 25, 26 and 30, the left side wall of thehopper has a vertical opening 88 and a pair of spaced outer flanges 89which extend outwardly from the left side wall of the hopper on oppositesides of the opening 88, as shown most clearly in FIG. 25. An upperhorizontal flange 83 extends outwardly from the left and rear side wallsof the hopper. The forwardmost flange 89 has an opening 84 just belowthe top flange 83, as shown in FIG. 25. Referring also to FIG. 25, apair of elongated reinforcing plates 82 are fastened to the outersurfaces of the outer flanges 89 by bolts 91. The bolts 91 are alsoutilized to fasten the hopper 87 to a pair of chain guide plates 90which are mounted to a hopper feeder support 92 which is, in turn,mounted on a base plate 93 by means of bolts 95. The chain guide plates90 are separated by a plurality of tubular spacers 97 through which thebolts 91 extend. A support bracket 94 is also mounted on the base plate93 and is fastened to the side of the hopper feeder support 92 as shownin FIG. 24. A support bar 96 is also mounted to the outside of the rearmost plate 90 by the bolts 91. A ball slide assembly 110 is mounted tothe support bar 96. A mixing bar mounting plate 111 is mounted to theball slide assembly 110. An endless conveyor chain 98 is located at thevertical side opening 88 and extends around a lower idler sprocket 101and an upper drive sprocket 100. The sprockets 100 and 101 are mountedon bushings 102 and are rotatively mounted on the hopper feeder support92. The upper drive sprocket 100 is driven by a stepper motor 103 whichis mounted on the support 92. One section of the conveyor chain 98 isguided along grooves in the outer longitudinal edges of the guide plate90 and is located between the inner surfaces of the flanges 89 whichdefine the opening 88. A plurality of spaced bars 99 are located on theoutside of the conveyor chain 98 and slant downwardly and forwardlytoward the event conveyor. The chain 98 travels upwardly from the bottomof the hopper 87 at an angle from the vertical. An idler sprocket shaft112 extends through the bushing 102 and rotates with the idler sprocket101, see FIGS. 26 and 27. The forward end of the shaft 112 is fixed to acam wheel 113 so that the cam wheel 113 rotates with the idler sprocket101 by means of a clamp 114. A lever arm 115 is pivotally mounted on ashaft 116 which is mounted in an adjusting fixture 117 which is locatedat a notch 118 in the left hand edge of the hopper feed support 92. Thepivoted end of the lever arm 115 has a flanged bearing 122 which enablesthe lever to pivot freely on the shaft 116. The opposite end of thelever arm 115 has a slot 121 which receives a pin 120 of a mixing block109. The mixing block 109 is fixed to the mixing block mounting plate111 and has an upper surface 123 which slants downwardly from back tofront at the same angle as the bars 99. The mixing block 109 is parallelwith the section of the conveyor 98 which travels upwardly along thevertical opening 88 of the hopper and is located adjacent the bars 99. Aball bearing follower 119 is rotatively mounted on the lever arm 115 andrides in a cam slot, not shown, on the rear side of the cam wheel 113.As the cam wheel 113 rotates with the idler sprocket 101, the lever arm115 oscillates about the shaft 116. The right hand end of the lever arm115, as viewed in FIG. 24, moves up and down and in turn causes themixing block 109 to move up and down. The timing of the upper movementof the block 109 is such that the block moves upwardly at the same rateas the upward movement of the conveyor chain 98. The cuvettes are storedin the hopper 87 in a random manner. The mixing block 109 serves twofunctions. The first function is to agitate the cuvettes within thehopper 87, and the second function is to assist in guiding the cuvettesonto the bars 99, one cuvette per bar. As the cuvettes are carriedupwardly by the bars 99, the ends of the cuvettes are guided by theinner surfaces of the flanges 89 to maintain the cuvettes in position onthe bars 99. As each cuvette reaches the opening 84, it slides forwardlyalong its respective bar 99 through the opening 84, see FIGS. 25 and 27,in the forwardmost flange 89 and falls into the orientation chute 131.

The orientation chute 131, as viewed in FIGS. 24, 27 and 30, consist ofa left hand plate 129 and a right hand plate 132 which are connectedtogether by screws 139 and held in a spaced parallel relationship by apair of spacer blocks 133. Each plate 132 and 129 has an upperslide-surface 134 which define, therebetween, a slot 135 toward theevent conveyor. The slide surfaces 134 end at a downward angle from backto front and at a downward angle toward the slot 135. As each cuvette 40falls through the opening 84 from the conveyor chain 98 to theorientation chute 131, the bottom end of the cuvette falls into the slot135 and the flanges 58 are supported on the slide surfaces 134. Thisenables the cuvette 40 to slide down the surfaces 134 in a nearlyupright orientation. The chute 131 is mounted to the hopper feedersupport 92 by a chute support bracket 130. A chute end plate 136 isattack to the front edges of the plates 129 and 132 by screws 137. Theplate 136 stops the downward slide of the cuvettes 40. The end plate 136has a hole 147 for receiving a position sensor 148 which is mounted on aPC board 138. The PC board 138 is mounted on the plate 136 by fasteners149. The forward end of each slide surface 134 has a flat upper surface127 for receiving a flat spring 128 which helps to insure that thecuvette remains in the slot 135 when the cuvette strikes the end plate136. The forward end of the slot 135 has a widened portion or accessopening 141 which is slightly greater in width than the distance betweenthe outer edges of flanges 58, see FIG. 30. The access opening 141between the plates 129 and 132 enables the cuvette to fall between theplates into the orientation tube 140. The cuvette falls between a pairof opposed guide surface 142 and 143 along the inwardly facing surfacesof the plates 129 and 132, respectively. The guide surface 143 has anupwardly facing jutting surface 144. The guide surface 142 has arecessed portion 145 which forms a downwardly facing undercut surface146. The undercut surface 146 is opposed to the jutting surface 144 ofthe plate 132. The orientation tube 140 has a top opening 150 and abottom opening 151 and extends from the bottom of the orientation chute131 to the top of the preheater section 38. When the cuvette falls intothe access opening 141 at the end of the orientation chute, one of theflanges 58 of the cuvette strikes the jutting surface 144. This deflectsthe cuvette laterally toward the recessed portion 145 of the left handplate 129. As the cuvette shifts laterally, the opposite flange of thecuvette strikes the recessed portion 145 just below the downwardlyfacing undercut surface 146. This traps the flange of the cuvette belowthe undercut portion 146 and prevents the cuvette from accidentallyflipping upside down when it reaches the end of the chute 131. Thecuvette, thereafter, falls in an upright orientation along the guidesurface 142 and 143 into the orientation tube 140 through the topopening 150 and through the bottom opening 151 into the preheatersection 38. The orientation tube 140 has a helical twist which causesthe cuvette to rotate approximately 90° about its vertical axis so thatwhen the cuvette falls into the preheater section 38, the broad sides 56of the cuvette are forward and back as well as the flanges 58.

Referring to FIG. 29, the preheater section 38 comprises a pair ofspaced horizontal bars 158 and 159 which define therebetween a verticalslot 160. Each of the bars 158 and 159 has a top edge 161. When acuvette falls from the bottom of the orientation tube 140, the body ofthe cuvette falls into the slot 160 and the flanges 58 rest on the topedges 161. Plunger 19 is moved to its extended position into the slot160 by the motor 25 from left to right as viewed in FIGS. 3, 32 and 33.The plunger 19 is moved from left to right a distance which isapproximately or slightly more than a cuvette width which pushes all ofthe cuvettes in the preheater section toward the cuvette dispense andincubation section 39. The plunger 19 is then retracted by the motor 25to allow a subsequent cuvette to fall from the orientation tube 140 intothe preheater section 38. The motor 25 is activated to separate theplunger 19 once every twenty seconds or when a test is requested. Thecuvettes are deposited into the orientation tube 140 at a faster ratethan they are pushed along the preheater section 38 so that the tube 140becomes full of cuvettes as generally shown in dotted lines in FIG. 29.The sensor 148 is a reflective object sensor which indicates thepresence of a stationary cuvette when the tube is full. The sensor 148forms part of the overall analyzer control system and is effective tostop the motor 103 when the sensor senses a stationary cuvette at thetop of the orientation tube. The software which is used to control theinstrument keeps track of the cuvettes as they are subsequently used outof the orientation tube and controls when the stepper motor 103 isreactivated. The preheater section 38 contains a thermistor forcontrolling a pair of solid state DC driven thermoelectric modules(TEMs) which maintain the temperature of the preheater section at a settemperature of 37° C. TEMs are also known as thermoelectric coolingcouples which are used to maintain a predetermined temperature bytransferring heat from one mass to another. The transfer of heat isreversed by reversing the direction of current flow. The machineframework provides a heat sink for the pre-heater section 38. When thetemperature of the pre-heater section is below the set temperature, heatis transferred from the machine framework to the pre-heater section 38.When the set temperature of the pre-heater section is above the settemperature, as detected by the thermistor, the current through the TEMsis reversed and heat is transferred from the pre-heater section 38 tothe machine framework. The cuvette dispense and incubation section 39 isalso provided with a thermistor at two spaced strategic locations. Eachthermistor controls a pair of thermoelectric modules (also strategicallyplaced) for maintaining the cuvette temperature at 37° C. throughout thechemistry event line. In the particular embodiment shown, the preheatersection 39 holds seventeen cuvettes and the cuvette dispense andincubation section 39 holds forty-five cuvettes.

Referring particularly to FIGS. 32 and 33, the track section 23 is shownin greater detail. The entire track section, including the preheatersection 38 and the dispense and incubation section 39, is covered by atop plate 162 which has a plurality of access openings at the dispensepoints 44, 45, 46 and 47. The plate 162 has an opening 186 at the sampledispense point 44 as shown in FIG. 33A. The plate 162 has openings 187and 188 for the reagent dispense points 45 and 46, respectively, asshown in FIG. 33B and an opening 189 for the reagent dispense point 47as shown in FIG. 33C.

Referring particularly to FIG. 32A, the plunger 19 (not shown) has a tab154 which extends horizontally toward the motor 25. When the plunger isin the outer or retracted position, it extends between a pair of spacedcomponents of an interruption sensor 155. The sensor 155 has a phototransmitting portion which directs a beam toward a photo receivingportion. When the beam is interrupted by the tab 154, a signal istransmitted to the CPU to indicate that the plunger is at the "home"position. (After a predetermined time period or when another test isrequested), the stepper motor 25 is actuated for a predetermined numberof steps to move the plunger 19 a predetermined distance out to theextended position. The motor is then reversed to bring the plunger backuntil the sensor 155 is interrupted by the tab 154 at the "home"position. All of the "interrupter" sensors described hereinafter areconnected to the CPU through the machine controller board and operate inthe same manner as the sensor 155. The cuvettes are pushed along thepreheater section 38 and into the cuvette dispense and incubationsection 39, at which point they are positively conveyed by a pair ofconveyor belts 167 and 168. Each of the conveyor belts 167 and 168 has aplurality of teeth 164 on one side of the belt for engaging the teeth 59of the cuvettes. A stepper motor 42 has a drive shaft 181 which isrotated in a clockwise direction when viewed from the front. The belt168 is driven by the motor 42 through the toothed drive pulley 170 whichis located between and below a pair of idler pulleys 171 and 179. Thebelt 168 extends over the pulley 179 to and around an idler pulley 178at the beginning of the incubation section 39. The belt 168 then travelsalong the front edge of the incubation section 39 to an idler pulley 172at the end of the section 39 and then back over the idler pulley 171 tothe drive pulley 170. The teeth 164 of the belt 168 face upwardly as thebelt 168 extends around the drive pulley 170 and the idler pulleys 171and 179 so that the teeth 164 of the belt engage the tee of the drivepulley 170. As the belt travels to the pulley 178, it gradually assumesa vertical orientation so that the teeth 164 face forwardly. As the beltextends around the pulley 178 and travels along the front edge of thedispense and incubation section 39, the teeth 164 face rearwardly and,thereby, engage the flanges 58 of the cuvettes. The belt 168 continuesin a vertical orientation around the idler pulley 172 and graduallyreassumes its horizontal orientation as it reaches the idler pulley 171.The pulleys 179 and 171 are rotatably mounted on horizontal shafts 182and 183, respectively. The pulleys 178 and 172 are rotatably mounted onvertical shafts 180 and 184, respectively. The drive belt 167 is locatedon the rear side of the dispense and incubation section 39 and is drivenlongitudinally by a drive pulley 175 which is fixed to the drive shaft181. The drive pulley 175 has external teeth 191 and is located betweenand below idler pulleys 174 and 176. The belt 167 extends over the idlerpulley 176 which is rotatively mounted on the horizontal shaft 182 andaround an idler pulley 177 which is rotatively mounted on a verticalshaft 190. The belt 167 then extends along the back side of the cuvettedispense and incubation section 39 to and around an idler pulley 173which is rotatively mounted on a vertical shaft 185. The belt 167 thenextends over the idler pulley 174 which is rotatively mounted on thehorizontal shaft 183 and back to the drive pulley 175. The belt 167 hasa plurality of the 193 on one side of the belt. The teeth 164 on thebelt 167 face upwardly as the belt 167 extends over the idler pulley 174and under the drive pulley 175 and back up around the idler pulley 176.The teeth 193 of the belt 167 are in drive engagement with the teeth 191of the drive pulley 175. When the belt 167 travels between the pulley176 and the pulley 177 it gradually assumes a vertical orientation sothat the teeth 193 face forwardly as the belt travels along theaspiration and incubation section 39 to the idler pulley 173. As theinner sections of the belts 167 and 168 travel from left to right asviewed in FIGS. 32 and 33, the rearwardly facing teeth of the belt 168and the forwardly facing teeth of the belt 167 engage the flanges 58 ofthe cuvettes 40 to advance the cuvettes along the event track ordispense and incubation section 39 for a predetermined time periodduring the twenty second system cycle.

Sample Transport System

The sample transport system consists of a sixty position sample tray forreceiving sample containers conning test samples, calibrators, controls,and diluents; a laser bar code reader; and a digital diluter. The sampletray consists of two concentric rings, each capable of holding a nixedpopulation of various tubes and sample containers. The outer ring canaccommodate thirty-four sample containers, the inner ring twenty-sixsample containers. Each position has a spring clip so that differentsizes of sample containers can be accommodated. The bar code readerrecognizes six versions of bar code language, and recognizes theidentity of each bar coded sample and the identity of the bar codedtray. The operator may program the analyzer to automatically repeat anysample whose initial test result exceeds a select range. Also, for mostassays, the system will automatically dilute and re-assay any sampleabove the range of the standard curve, if desired. Various dilutionratios are selectable, based upon sample size. The sample aspirating anddispensing probe is specially coated and has capacitance level sensingin order to recognize the surface of the sample. This insures thatliquid is present in a sample container before aspirating, as well asminimizing immersion into the test sample. After each aspiration anddispensing cycle, the inner and outer surfaces of the probe arethoroughly washed with deionized water at a wash station to minimizesample carryover.

The sample transport system 26 is shown in FIGS. 36-42. Referring firstto FIGS. 38, 39 and 41, the transport system 26 includes a fixed basewhich is generally indicated by the reference numeral 211 and which ismounted in a fixed position on the machine framework in front of thecuvette dispense and incubation section 39. The fixed base 211 includesan upper horizontal plate 212 and three descending legs 213, each with ahorizontally and outwardly extending foot portion 214. Each foot portion214 supports a roller 247 which is rotatively mounted on a horizontalshaft 215 for rotation about a horizontal axis. Each foot 214 alsosupports a roller 218 which is rotatively mounted on a vertical shaft217 for rotation about a vertical axis. An electric stepper motor 219 isfixed to the bottom of the upper plate 212 and has a drive shaft 220which extends through a hole 216 in the upper plate 212. A frictiondrive wheel 221 is fixed to the outer end of the shaft 220 for rotationtherewith. An inner tray, generally indicated by the reference numeral222, and an outer tray, generally indicated by the reference numeral223, are rotatively mounted on the base 211 for rotation independentlyof one another about a vertical axis 209.

The inner tray 222 includes an inner hub portion 225 which is relativelymounted on a vertical shaft 224 which is fixed to the upper plate 212and which extends along the vertical axis 209, see FIG. 38. The innerhub portion 225 has a downwardly ending annular flange 226 which is infrictional engagement with the drive wheel 221. When the motor 219 isactuated, the drive wheel 221 is rotated bar the shaft 220 which, inturn, rotates the inner hub portion 225 about the axis 209 due to thefrictional engagement of the roller 221 against the inner surface of theannular flange 226. The inner hub 225 has an outwardly extendingcircular flange 208 at the bottom of the hub. The flange 208 isrotatably supported on the rollers 247. The inner tray 222 also includesan outer hub 227 which has an outer annular flange 228 which supports aplurality of receptacles 229 for supporting a plurality of samplecontainers, see FIG. 37. The receptacles 229 are arranged in an innercircle which is concentric with the axis 209. Each receptacle 229 has anoutwardly facing opening 195.

The outer tray 223 includes a drive ring 230 which has an outerdownwardly extending annular flange 231. The annular flange 231 has aninwardly facing annular groove 232 for receiving the rollers 218 whichsupport the drive ring 230 for rotation about the axis 209. The drivering 230 supports an outer ring 233 which contains a plurality ofupwardly extending receptacles 234 for supporting a plurality of samplecontainers. The receptacles 234 are arranged in an outer circle which isconcentric with the axis 209 and is located outside of the circle ofreceptacles 229 as shown in FIG. 37. Each receptacle 234 has anoutwardly facing opening 260. Each of the receptacles 229 and 234 is atleast partially lined with a metal plate 270 which has a plurality ofinwardly protruding resilient fingers 271. The fingers provide a snugfit for a test tube or sample container and enable test tubes ofdifferent diameters to be used and held securely. The plates 270 andfingers 271 also provide a ground connection to the metallic machineframework to provide one component of a capacitance level sensing systemto be described in a later section entitled: "SAMPLE PROBE TRANSPORTSYSTEM". The outer tray 223 is rotated independently of the inner tray222 by means of a stepper motor 235 which is fixed to a mounting plate236 which is, in turn, supported on the framework of the machine. Thestepper motor 235 has a drive shaft 237 which is fixed to a drive pulley238. A pulley 239 is fixed to a vertical shaft 241 which is mounted forrotation on the plate 236. The pulley 239 is driven from the pulley 238by a timing belt 240. A drive wheel 242 is fixed to the pulley 239 andis in frictional engagement with an outer surface of the flange 231.When the motor 235 is activated, the roller 242 is rotated about theaxis of the shaft 241 which, through its frictional engagement with theouter surface of the flange 231, causes the drive ring 230 to rotateabout the axis 209. This rotation is totally independent of the rotationof the inner tray 222 by the stepper motor 219.

Referring to FIGS. 40 and 42, a PC board 245 is mounted to the machinebase adjacent the sample transport system 26. The PC board 245 supportsa plurality of interrupt sensors for the inner and outer trays. Thesensors are arranged in two groups, an outer group for the outer ring,and an inner group for the inner ring. The outer group includes a pairof spaced outer sensors 246 and an inner home sensor 266. The innergroup includes a pair of inner sensors 244 and an inner home sensor 267.The outer ring 230 has a single downwardly descending home tab 253 whichinterrupts the beam of the home sensor 266 to determine a startingposition for the outer ring at the beginning of a test or a series oftests A plural of tabs 268 extend downwardly from the drive ring 230 ofthe outer tray 223 outside of the home tab 253 and extend in a circleabout the axis 209. As the outer ring rotates about the axis 209, thetabs 268 pass through both sets of sensors 246. There is a tab 268 foreach sample position of the ring 230 so that each time that the ring isrotated one position, the beam in each of the sensors 246 is interruptedto provide a signal to the CPU to indicate that the outer tray 223 hasmoved one position. The distance between the two sensors 246 differsfrom the spacing between two adjacent tabs 268 so that the sensors arenot interrupted simultaneously. This enables the control electronics todetermine the direction of rotation of the ring 230. To position aparticular bottle or sample container about the axis 209, a command isgiven to the stepper motor 235 to move a number of steps in a certaindirection and acceleration. The optical interrupt sensors 246 count thenumber of positions moved by the drive ring 230 to determine the finaldesired position of the ring. When the correct number of transitionshave occurred, the stepper motor 235 will move a calibrated number ofsteps past the transition point and stop. This will be the finalcontainer positioning point. The CPU is programmed to move the ring 230and outer tray 223 in whichever direction will result in the smallestamount of rotation of the ring for each new sample container position. Asingle "home" tab 269 extends downwardly from the inner tray 222 forinterrupting the beam of the home sensor 267 to determine the startingor "home" position of the inner tray. A plurality of tabs 243 extenddownwardly from the tray 222 outside of the "home" tab 269 and extend ina circle which concentric with the axis 209. The tabs 243 interact withthe interrupt sensors 244 for controlling the stepper motor 219 andselectively positioning the inner tray 222 in the same manner as thetabs 268 and sensors 246 are utilized to selectively position the outertray 223. The inner and outer trays are moved selectively andindependently to position a specified predetermined sample container toa predetermined pickup position for aspiration by the sample aspiratingand dispensing probe 24. Referring to FIG. 22, the pickup position forthe outer tray is located at the opening 255 in the outer cover 257. Thepickup position for the inner tray is located at the opening 256 in theouter cover 257. A bar code label is affixed to the outer wall of eachsample container. The label has a specific bar code which identifies thetest sample within the container. All of the information relating to thesample, such as the name of the patient and the tests which are to beperformed with the sample, are stored within the memory of the centralprocessing unit. Referring to FIG. 22, a bar code reader 258 is locatedadjacent the sample transport system 26 and has two lines of sight whichare indicated by the dotted lines 259 and 272. Prior to a run of tests,the receptacles in the inner and outer trays are charged with samplecontainers each containing its own specific bar code which can be viewedthrough the openings 260 in the outer parts of the receptacles 234 andthe clear plastic cover 257. The outer tray 223 is rotated about theaxis 209 so that each sample container passes through the lines of sight272 and 259 relative to the bar code reader 258 so that the bar code oneach sample container can be read by the bar code reader. The energybeam from the transmitting portion of the bar code reader 258 passesalong the line of sight 272 and the beam is reflected back from the barcode label on the sample container along the line of sight 259 to thebeam receiving portion of the bar code reader. The vertical openings 260and the transparency of the outer cover 257 enable the bar codes on thesamples to be "seen" by the bar code reader. This enables the identityof each sample container to be correlated with the position of the outertray relative to a home position. After all of the sample containershave been read by the bar code reader, the outer tray 223 is positionedso that a gap 261 in the circle of receptacles 234 is aligned with thelines of sight 259 and 272. This enables the bar codes on the samplecontainers in the inner tray 222 to be exposed through openings 195 inthe outer portions of the receptacles 229 to the bar code reader 258.The inner tray 222 is rotated so that each sample container in the innertray passes through the lines of sight 259 and 272 so that the specificbar code of each sample in the inner tray 222 is read by the bar codereader. This information is utilized by the central processing unit tocorrelate the position of each sample container in the inner tray 222relative to the home position of the inner tray.

Referring particularly to FIGS. 39 and 41, a contact ring 250 isfastened to the drive ring 230 by a screw 262 which also mounts apositioning key 263 to the drive ring 230. A contact ring 252 isfastened to the upper wall of the hub 225 by a screw 264. Positioningkey 265 is fixed to the hub 225 at the base of the flange 226. The metalgrounding wire 248 is connected to the contact ring 252 and connected tothe keys 265 and 263 by a connecting wire 249. These elements form partof the grounding system for grounding the fingers 271 to the machineframework.

The bar code-labeled sample containers may be loaded in any order in thesample tray. The analyzer will read all bar codes automatically, andidentify the sample and its position in the tray. If bar code labels arenot used, a worklist printout is utilized, which directs placement ofsamples in specific sample tray positions.

Reagent Transport System

The reagent transport system or tray provides a carrier for twenty-sixreagent bottles or containers, sufficient for up to thirteen differentassays. The inner portion is made to specifically accept the solid-phasereagent containers, and periodically agitates these containers tomaintain homogeneity of the solid phase reagent. This mixing action isaided by the design of the reagent bottles, which have agitator finsmolded into their inner walls. The tracer or labeled reagent bottles arealso specially shaped to automatically orient the identifying bar codelabel affixed to the container, and are loaded into the outer positionson the reagent tray. Reagents are bar code labeled. A reagent laser barcode reader records the loaded position of each specific reagent,including identity and lot number, making random loading permissible.Reagents may be loaded directly from refrigerated storage, since theyare warmed to 37° C. before dispensing. The three reagent aspirating anddispensing probes have capacitance level sensing and may be programmedto make an initial reagent level check before starting an assay run toinsure that adequate reagent volumes have been loaded to complete thescheduled worklist stored in the CPU. Reagent volumes used range from50-450 uL, depending on the assay, and specific reagents may be added tothe ample in the cuvette by each of the three reagent probes, withincubation time of 2.5 to 7.5 minutes, depending on optimal conditionfor specific assays. Reagent probes, like the sample probes, arethoroughly washed with deionized water between dispensings.

Referring to FIGS. 43-49, the reagent transport system is generallyindicated by the reference numeral 27. The reagent transport system 27comprises a fixed supporting base 286 which is fixed to the machineframework 283 and an electric stepper motor 287 which is fixed to thesupporting base 286 by fasteners 282 and connecting rods 285. Thestepper motor 287 has a drive shaft 290 which is fixed to a motor hub291 by a trantorque clamp 280. The drive shaft 290 is rotated about avertical drive axis 293. The base of the motor hub 291 consists of aring of upwardly facing gear teeth 292. The circular spill tray 288 hasa central circular opening 289 and is fixed to the supporting base 286by a plurality of fasteners 279 so that the stepper motor 287 extendsupwardly through the opening 289. Referring to FIGS. 45 and 46, asupport ring 294 is located concentrically of the central vertical axis293 and has a central circular opening 295 and a plurality of smalleropenings 308 which are arranged in a circle which is concentric with theaxis 293. A reagent tray 296 is mounted on the support ring 294 andcontains a ring of inner pockets 297 and a ring of outer pockets 299.The pockets 297 and 299 are arranged in inner and outer concentriccircles, respectively, about the axis 293. Each outer pocket 299contains a tubular outer bottle or reagent container holder 298 which isfixed to the pocket by a fastening disc 301. The connector 301 extendsthrough an aperture 302 at the base of the pocket to the support ring294 for fastening the reagent tray 296 to the ring 294. When a container60 of labeled or tracer reagent is placed in the pocket 299, the tubularholder 298 extends between the skirt 63 and the main body portion 64 asshown in FIG. 45.

Each inner pocket 297 contains an inner container holder 300. Afastening disc 303 bears against the bottom wall of the holder 300 andhas a vertical shaft 304 which extends through an opening in the bottomwall of the holder. The fastening discs 301 and 303 are metallic and aregrounded to the machine framework. The discs 301 and 303 provide onecomponent of a capacitance level sensing system which is described in afollowing section entitled "REAGENT PROBE TRANSPORT SYSTEM". A gear 306is fastened to the bottom of the holder 300 by a pair of screws 305which also effectively clamp the fastening disc 303 and the gear 306against the bottom wall of the holder 300. The bottom of the shaft 304extends below the gear 306 and into a pair of flanged bearings 307 whichare mounted in one of the apertures 308 of the support ring 294. Thisenables each holder 300 and its respective gear 306 to rotate about itsown central longitudinal secondary axis 278. The gears 306 extend abouta ring gear 309 and are in driving engagement with the outer teeth ofthe ring gear, see FIG. 46. The ring gear 309 has a large centralopening 277. A pair of pins 310 are fixed to the gear 309 and end belowthe gear into driving engagement with the teeth of the ring gear 292,see FIG. 45. Actuation of the stepper motor 287 causes the hub 291 inthe ring gear 292 to rotate about the axis 293. This causes rotation ofthe ring gear 309 through the drive pins 310. The ring gear 309, inturn, drives all of the satellite gears 306 for rotating each bottleholder 300 about its respective secondary axis 278. The ring gear 309 isfully supported by the satellite gears 306. A plurality of retainers 311are fixed to the ring gear 309 and extend below the gear 309 forstraddling the inner edge of the support ring 294. The bottle holder 300holds a solid phase bottle or reagent container 75. The side walls ofthe holder 300 has a plurality of vertical slots 276 which form aplurality of resilient fingers 274 which extend between the main body 76and the skirt 80 of the reagent bottle or reagent container 75 forholding the reagent container 75 in a friction fit. The step motor 287is reversible and controlled by the central processing unit to oscillatethe drive shaft 290 at predetermined intervals. Each of the bottleholders 300 is adapted to receive a solid phase reagent container 75.The oscillations of the holder 300 provide the necessary motion to thereagent container 75 for enabling the fins 81 to agitate the solid phasereagent solution within the bottle 75 and, thereby, maintain a uniformconcentration of the solid phase elements within the solution. Each ofthe bottle holders 298 is adapted to receive a labeled reagent container60 which does not require agitation. Referring particularly to FIGS. 45and 47, a ring gear 312 encircles the spill tray 288 and is mounted forrotation on the supporting base 286 about the axis 293. The lower partof ring gear 312 has an inwardly facing V-shaped bead 275 which engagesa plurality of V-guide wheels 323 which support the ring 312 forrotation about the axis 293. Each wheel 323 is rotatively mounted on avertical shaft 324 which is fixed to the base 286. The ring gear 312supports the support ring 294 and the reagent tray 296. Referring alsoto FIGS. 48 and 49, part of the ring gear 312 has an annular flangewhich is opposite the V-shaped beads 275 and contains a ring ofoutwardly facing gear teeth 329 which are in driving engagement with anidler gear 319 which is keyed to a vertical shaft 320. The shaft 320 isrotatively mounted in flanged bearings 321 which are supported onflanges 322 of a motor mount 314. The motor mount 314 has a circularbore 316 which contains a drive gear 318 which is fixed to the driveshaft 317 of a stepper motor 315. The stepper motor 315 is fixed to themotor mount 314. The wall of the bore 316 of the motor mount 314 has alateral opening which enables the drive gear 318 to engage the idlergear 319. Actuation of the motor 315 causes the drive gear 318 to drivethe ring gear 312 through the idler gear 318 about the vertical axis293. The inner and outer pockets 297 and 299, respectively, are enclosedwithin a clear stationary plastic covers 327. The cover 327 has aplurality of openings 328, 338, 339, 340, 341, and 342 which provideaccess to the bottles within the pockets 297 and 299 by reagentaspirating and dispensing probes to be described in a later section, seeFIG. 22.

Referring to FIG. 47, a PC board 330 contains a pair of interruptersensors 331 and 336 and a photo reflector sensor, not shown, which islocated beneath the sensors 331 and 336. The optical reflector sensorhas a beam transmitting portion and beam receiving portion. If a beamfrom the transmitting portion strikes a reflective surface, the beam isreflected back to the receiving portion of the sensor. When the beam isnot reflected back, the sensor generates a signal to the CPU. The PCboard 330 is mounted to the base plate 286 so that the sensor opticalreflector face outwardly toward the ring 312. The beam from thetransmitting portion of the beam reflector sensor strikes the ring 312and is reflected back to the beam receiving portion of the sensor. Thering 312 has an aperture 326, see FIG. 49, which is at the same level asthe beam from the photo reflector sensor. At the beginning of a testingsequence, the ring 312 is rotated about the axis 293 until the beam ofthe photo reflector sensor is aligned with the aperture 326. When thisoccurs, the beam passes through the aperture and is not reflected backto the sensor. The absence of the reflected beam initiates a signal tothe CPU to indicate the "home" or starting position of the reagent trayat the beginning of a series of tests. Referring to FIG. 47, the ring312 has a plurality of tabs 334 which extend inwardly from the ring 312and which pass between the two spaced elements of each interruptersensor 331 and 336 for interrupting a beam from each optical sensorwhich provides feedback to the control electronics for reagent bottlepositioning. There is a tab for each reagent bottle position in the tray296 so that each time that the ring is rotated one position, the beam ineach of the sensors 331 and 336 is interrpted to provide a signal to theCPU to indicate that the tray has moved one position. The distancebetween the two sensors is less than the spacing between two adjacenttabs 334 so that the sensors 331 and 336 are not interruptedsimultaneously. This enables the CPU to determine the direction ofrotation of the reagent tray. To position a particular bottle orcontainer to a reagent probe pickup or aspiration position, a command isgiven to the stepper motor 315 to move a fixed number of steps in acertain direction. This causes the reagent tray 296 to rotate along withthe tabs at the bottom of the drive ring 312. The sensors 331 and 336counts the number of tab transitions and determines the position of thereagent tray 296. When the correct number of transitions have occurred,the stepper motor 315 will move a calibrated number of steps past thetransition point and stop. The bottle containing the designated reagentwill thereby be positioned at the predetermined pickup point for one ofthe reagent probes.

A photo reflective sensor 337 is mounted on the plate 286 and directs alight beam upwardly. The motor hub 291 has a bottom reflective surfacewhich has a plurality of spaced apertures. As the hub 291 oscillates,the beam from the sensor 337 is alternately reflected back to the sensorby the bottom reflect surface of the hub and absorbed by the aperturesin the bottom surface. This provides appropriate signals to the CPU toindicate that the hub is being oscillated at predetermined intervals.

Each reagent container has a bar code label affixed to its outer skirtportion. The label contains a specific bar code which identifies thereagent within the container. The information relating to all of thereagent in the bar codes associated with the reagents are stored withinthe memory of the central processing unit. Referring to FIGS. 43 and 22,a bar code reader 332 is located adjacent the reagent transport system27. The bar code reader 332 transmits an energy beam along a line ofsight which is indicated by the dotted line 333. The beam is reflectedback go the bar code reader 332 from the bar code label along a line ofsight which is indicated by the dotted line 344. The return beam alongthe line of sight 344 is received by the beam receiving portion of thebar code reader. The bar code in the preferred embodiment is printed onthe label for each reagent bottle in a vertical direction. The innerpockets 297 and outer pockets 299 are staggered with respect to eachother. As the reagent tray 27 is rotated about the axis 293 by thestepper motor 315, the inner and outer pockets alternately pass throughthe lines of sight 333 and 334 of the bar code reader 332. The steppermotor 287 is also utilized during the initial reading of reagentcontainer bar codes prior to a run of tests Referring to FIGS. 43 and46, there is a relatively large space between each outer pocket 299.Each inner pocket 297 is horizontally aligned with the space between twoadjacent pockets 299. A vertical wall 335 which separates the inner andouter pockets 297 and 299, respectively, has a relatively large opening328 at each space between outer pockets 299 so that each reagentcontainer is exposed to the line of sight of the bar code reader whenthe container is rotated about the axis 293 by the stepper motor 315. Asthe reagent tray 27 is rotated about the axis 293, each reagentcontainer or bottle in the ring of inner pockets 297 is given one andone-half revolutions per pass of a reagent container 75 through thelines of sight 333 and 334 to insure that the bar code is exposed to thereader. The bar codes on the bottles in the inner and outer pockets canbe read by the bar code reader 332 through the clear plastic cover 327.

The operator loads required assay reagents, in original bar code-labeledbottles, into the reagent tray in any order; solid-phase reagents on theinner bottle holders 300, labeled or tracer reagents on the outer bottleholders 298. Due to the design of the reagent bottles, it is notpossible to mis-load reagents. The analyzer will read all bar codesbefore initiating a run, identifying each reagent, its position, its lotnumber and expiration date. If greater than 50 tests of a specific assayhas been requested in the worklist, multiple bottles of the necessaryreagents may be loaded on the reagent tray and the analyzer will accessthem sequentially, as needed.

Sample Probe Transport System

Referring to FIGS. 50-59 and first to FIGS. 54 and 55, the sample probetransport system 24 comprises a fixed upper horizontal support plate357, and a sample probe supporting carnage, generally indicated by thereference numeral 363, which is mounted for horizontal back and forthmovement relative to the supporting plate 357. The support plate 357 hasan opening 366. A PC board 358 is fixed to the upper surface of theplate 357 by screws 359. The under surface of the PC board has aplurality of electrical junctions J1, J2, J3, J4 and J5 which extendinto the opening 366. A vertical bracket 364 is fixed to the undersideof the plate 357 at the rear end of the plate. An electrical steppermotor 365 is fixed to the forward side of the bracket 364 and has adrive shaft 369 which is rotatable about a horizontal axis. A lead screw371 is fixed to the drive shaft 369 through a drive coupling 370 andextends through a roll nut 409 which is fixed within a bore 408 of ablock 372. (See also FIG. 58.) The block 372 is mounted in a yoke 373between a pair of upper and lower dowel pins 374. The dowel pins 374enable the block 372 to pivot about a vertical axis to compensate forslight misalignments between the block 372 and a lead screw 371. Theblock 372 has a laterally extending horizontal shaft 375 which ismounted to the carriage 363 in a manner described herein below.

A guide bracket 360 is fixed to the underside of the plate 357 by thescrews 359 and has a downwardly facing horizontal groove 361. A carriagesupporting bar 362 is slidably mounted in the groove 361. The carriage363 is fixed to the sliding bar 362 by a screw 391 and an anti pivot rod387 which has a threaded upper end. The carriage 363 includes aforwardly facing vertical wall 376, a top horizontal wall 377 and alower horizontal wall 378. The top wall 377 has an aperture 389 and thebottom wall 378 has an aperture 388. The anti pivot rod 387 extendsfreely through the apertures 388 and 389 and is threaded into the block362. Referring also to FIG. 56, the wall 376 has a horizontal bore 379which has a bearing 380 at each end of the bore. The shaft 375 of theyoke 373 extends through the bore 379 within the bearings 380. Avertical lead screw 385 is rotatably mounted in upper and lower bearings383 and 384, respectively, in the upper and lower walls 377 and 378,respectively. The lower end of the lead screw 385 extends below thebottom wall 378 and is fixed to a pulley 386. An electrical steppermotor 394 is fixed to the underside of a rearwardly extending horizontalflange 393 of the carriage 363. The stepper motor 394 has a verticaldrive shaft 395 which is fixed to a pulley 396, see also FIG. 57. Thepulley 396 is drivingly connected to the pulley 386 through a timingbelt 397. The inner surface of the timing belt 397 has a plurality ofteeth for engaging corresponding teeth on the drive pulleys 396 and 386,(teeth not shown). A lead screw follower 401 is positioned between thewalls 377 and 378 and has a vertical bore 403 and a vertical bore 404which contains a roll nut 405 (see also FIG. 59). The anti pivot rod 387extends freely through the bore 403 and the lead screw 385 extendsthrough the roll nut 405. The roll nut 405 is fixed relative to thefollower 401 so that as the lead screw 385 is rotated about its verticalaxis, the follower 401 moves along the central longitudinal axis of thelead screw 385 relative to the walls 377 and 378. A probe holding arm402 is fixed to the forward end of the follower 401 and carries anaspirating and dispensing sample probe 407.

A PC board 398 is fixed to the carriage 363 and has an electricalconnector 399 which is connected to the electrical junction J2. Thestepper motor 394 has a connector 400 which is connected to theelectrical junction J4. The stepper motor 365 has a connector 368 whichis connected to the junction J5. The probe supporting arm 402 has a PCboard 406 which is connected to a connector 411 through a flexibleribbon 421. The connector is connected to junction 420 of the PC board398.

The stepper motor 365 is reversible. When the lead screw 371 is rotatedin one direction, the carriage 363 moves rearwardly along the centrallongitudinal axis of the lead screw 371 toward the flat bracket 364.This causes the carriage 363 and the sample probe 407 to move from aforward position to a rearward position relative to the sample tray.When the stepper motor 365 is reversed, the lead screw 371 is rotated inthe opposite direction. This causes the carriage 363 to move forwardlyand, thereby, move the sample probe 407 from its rearward position toone of two forward pickup positions above the sample tray. The sampleprobe 407 can also be positioned in intermediate positions betweenrearward and forward positions, as for example, above the wash station18. The motor 394 is also reversible. Rotation of the lead screw 385 inone direction causes the follower 401 and the arm 402 to move upwardly.Rotation of the lead screw 385 in the opposite direction, causes thefollower 401 and the arm 402 to move downwardly. The sample aspiratingand dispensing probe 407 is moved forwardly when it is in the upperposition until it reaches one of the sample pickup or aspirationpositions above the sample tray and is then moved downwardly to pick upa volume of a sample. The probe 407 is then moved to the upper positionand returned to a point above the wash station, whereupon it is moveddownwardly again for a wash cycle, or to its rearward position above oneof the cuvettes, whereupon it is lowered into the cuvette for depositingthe sample volume into the cuvette. The stepper motors 394 and 365 arecapable of making very precise step-by-step motions for very precisehorizontal and vertical positioning of the sample probe 407.

Referring to FIGS. 54 and 56, a plurality of spaced tabs 410 extendupwardly from the carriage 363 from front to back on one side of thecarriage. A single "home" tab 415 extends upwardly from the carriage 363on the opposite side of the carriage. When the carriage 363 reaches itsrearward "home" position, the tab 415 passes between the elements of aninterrupt sensor 413 which extends downwardly from the support plate357. The tab 415 interrupts a light beam between the two elements of thesensor 413 which initiates a signal to the CPU that the carriage hasreached its "home" position and the sample probe 407 is directly above acuvette at the sample dispense point 44. The upper portion of the probecarding arm 401 is determined by an interrupt sensor 416 which is fixedto the PC board 398. The PC board is fixed to the carriage 363 so thatit extends horizontally toward the probe carrying arm 401, see FIGS. 50and 56. The follower 401 has a tab 355 which extends toward the sensor416. The tab 355 cannot be seen in FIGS. 54 and 56 since it is locatedon the hidden side of the follower 401, but is indicated by dotted linesin FIG. 53. When the follower 401 reaches the upper position, the tab355 passes between the two elements of the sensor 416 and interrupts alight beam. The interruption of the light beam provides a signal to theCPU to indicate that the follower 401 and the probe 407 have reached theupper position. This insures that the carriage 363 can be safely movedto a new horizontal position at a predetermined point of time in theoperating cycle, whereupon the motor 365 is given pulses for apredetermined number of half steps. At the appropriate time, the motor394 is activated to move the arm 401 and the probe 407 downwardly. Foreach sample pickup cycle, the motor 365 is actuated for a predeterminednumber of half steps to move the carriage forwardly with the probe 407in the upper position from the home position until the probe 407 isabove the wash station 18. The motor 394 is actuator for a predeterminednumber of half steps to lower the probe 407 into the wash station 18 fora wash cycle. The probe 407 is then raised by reversing the steppermotor 394 for a predetermined number of half steps. The motor 365 isactuated for a predetermined number of half steps to move the carriage363 forwardly until the probe 407 is above the opening 255 or theopening 256 in the outer cover 257 of the sample transport system. Themotor 394 is actuated to move the follower 401, together with the arm402 downwardly to lower the probe 407 into the sample container which islocated beneath whichever of the openings 256 or 255 which is verticallyaligned with the probe 407. The lower position of the sample probe 407is determined by a capacitance fluid sensing system. The capacitancefluid sensing is a function of a signal change occurring through twoconductive materials such as the metal probe 407 and ground fluid andone non-conductive material such as air or plastic/glass samplecontainer. When the probe is in the upper position, the probe'sreference current is measured, as the probe moves downwardly seekingfluid, an increase in signal indicates the presence of fluid. When fluidis detected, the motor 394 is actuated for a predetermined number ofhalf steps to move the probe 407 a predetermined distance below themeniscus of the fluid. This distance is determined by the amount offluid to be aspirated, a large volume requiring a deeper penetration ofthe probe than a smaller volume. After aspiration of a volume of sampleby the probe 407, the probe is raised to its upper position, whereuponthe motor 365 is actuated for a predetermined number of half steps tomove the carriage 363 rearwardly to its "home" position so that theprobe 407 is directly above the sample dispense point 44. The motor 394is actuated for a predetermined number of half steps to lower the probe407 in the cuvette which is located beneath the dispense point 44. Thequantity of sample is then dispensed by the probe 407 into the curve.The probe 407 is raised to its upper position to begin another cycle. Asthe carriage moves between the "home" and forward positions, the tabs410 pass between the elements of an interrupt sensor 412. The tabs 410are positioned so that when the carriage stops at a forward position fora sample pickup or a wash cycle, none of the tabs 410 will interrupt thelight beam which passes from one element of the sensor 412 to the other.The light beam will pass through one of the spaces between the tabs 410or outside of the outer edge of one of the tabs when the probe isproperly positioned. If the probe is not properly positioned, due to amalfunction in the system, one of the tabs 410 will interrupt the lightbeam and a signal will be sent to the CPU to stop the machine. This willprevent the lowering of an improperly positioned probe and subsequentbreaking of the probe.

For most test protocols, the sample probe will make one forward stopafter the wash cycle to pick up a volume of sample from either the outertray or the inner tray. In some cases, the sample probe stops at both ofthe openings 255 and 256 to pick up a volume of diluent as well as avolume of sample. The diluent is generally a protein based solutionwhich is used to dilute a patient sample when an original test result isbeyond a test curve range. The type of diluent used should correspond tothe type of assay being performed by the analyzer. Diluent solutions arenormally placed in the inner tray. The sample probe picks up the diluentbefore picking up the test sample as to avoid contaminating the diluentwith sample. Other treatment liquid materials which are sometimes pickedup with a sample solution are pretreatment agents and releasing agents.A releasing agent is sometimes mixed with the sample for the purpose ofseparating the analyte from another molecule and rendering it availablefor reaction. A pre-treatment agent is a solution which is mixed andincubated with the test sample to protect the analyte from a releasingagent.

Reagent Probe Transport System

The reagent probe transport system is shown in FIGS. 60-72. Referringfirst to FIGS. 60-63, the reagent probe tort system is generallyindicated by the reference numeral 440 and includes the reagent probetransport systems R1, R2 and R3. The system 440 comprises an upperhorizontal support plate 441 which has openings 442, 443, 444 and 445. APC board 446 is fixed to the upper surface of the plate 441 and has aplurality of interrupter sensors on the undersurface of the PC boardwhich extend into the openings 442, 443, 444 and 445. Interruptersensors 448, 449, 450 and 451 extend into the opening 442. Interruptersensor 452 extends into the opening 443. Interrupter sensor 453 extendsinto the opening 444 and interrupter sensors 454 and 453 extend into theopening 445. A plurality of electrical junctions are also mounted on theother side of the PC board 446 and are accessible through the openings442, 443, 444 and 445. Junctions J11 and J12 are accessible through theopening 442. The junctions J13, J14 and J15 are accessible through theopening 443. Junctions J16, J17, J18 and J19 are accessible through theopening 444. Junctions 120, 121 and 122 are accessible through theopening 445. Three horizontal guide brackets 455, 457 and 459 are fixedto the underside of the support plate 441. The guide brackets 455, 457and 459 have elongated horizontal grooves 456, 458 and 460,respectively. Elongated carriage supporting guide bars 461, 462 and 463are slidably mounted in the grooves 456, 458 and 460, respectively. Theguide bar 461 is fixed to a reagent probe supporting carriage which isgenerally indicated by the reference numeral 464 and which forms part ofthe reagent probe transport system R1. The carriage supporting slide bar462 is fixed to a reagent probe supporting carriage which is generallyindicated by the reference numeral 465 and which forms part of thereagent probe transport system R2. The carriage supporting slide bar 463is fixed to a reagent probe supporting carriage which is generallyindicated by the reference numeral 466 and which forms part of thereagent probe sport system R3. Slide bars 461, 462 and 463 enable thecarriages 464, 465 and 466 to move forwardly and rearwardly relative tothe support plate 441.

A flat vertical rear bracket 467 is fixed to the back end of the supportplate 441 and extends downwardly from the under surface of the supportplate. A plurality of stepper motors 468, 469, 470 and 471 are fixed tothe front side of the plate 467. The stepper motors 468, 469, 470 and471 have forwardly extending and horizontal drive shafts 472, 473, 474and 475, respectively. The motors 468, 469, 470 and 471 have electricalconnectors 476, 477, 478 and 479, respectively, which are connected tothe electrical junctions J10, J12, J20 and J18, respectively, on the PCboard 446. A bracket 480 is connected to the right side of the supportplate 441 as viewed in FIG. 63 and fixedly supports a horizontal slidebar 481 which is slidably mounted in the horizontal groove 482 of aguide bracket 483. The guide bracket 483 is fixed to a guide rail 487which is fixed to the framework of the machine. A horizontally extendingslide bar 484 is fixed to the left side of the support plate 441 asviewed in FIG. 63 and is slidably mounted in a horizontal groove 485 ina guide bracket 486. The guide bracket 486 is fixed to an upwardlyextending arm of a U-shaped bracket 488 which is fixed to a guide rail489. The guide rail 489 is, in turn, fixed to the machine framework.Brackets 483 and 486 are fixed relative to the machine frame and theslide bars 484 and 481 are fixed to the support plate 441. The supportplate 441 is able to move forwardly and rearwardly between the guidebrackets 486 and 483, along with the carriages 464, 465 and 466 whichare supported from the underside of the support plate 441.

The forward and backward motion of the support plate 441 is provided bythe stepper motor 469. The drive shaft 473 of the motor 469 is fixed toa horizontally extending lead screw 490 through a coupling 491 (See alsoFIG. 67). The lead screw 490 extends through a roll nut 497 which islocated in a bore 492 of a block 493. The block 493 is pivotally mountedbetween the parallel arms of a yoke 494 by means of a pair of upper andlower dowel pins 495 which end into a bore 435 of the block 493. Theroll nut 497 is fixed to the block 493 so that as the lead screw 490 isrotated, the block 493 moves along the central longitudinal axis of thelead screw. The pivoting motion of the block 493 along the longitudinalaxis of the bore 435 within the yoke 494 compensates for any possiblemisalignments between the block 493 and the lead screw 490. The yoke 494has a shaft 496 which extends upwardly through a tubular follower guide437 which is located in an aperture 439 in a bottom wall 438 of theU-shaped bracket 488, see FIG. 63. The shaft 496 rides in a pair ofbearings 436 at opposite ends of the follower guide 437. When the leadscrew 490 is rotated upon actuation of the motor 469, there is relativemotion between the block 493 and the lead screw 490 along thelongitudinal axis of the lead screw. Since the block 493 is fixedrelative to the machine framework this motion causes the lead screw 490and the motor 469 to move relative to the machine framework which, inturn, causes the support plate 441 to move forwardly or backwardly,depending upon the rotation of the lead screw 490.

The forward position of the plate 441 is the normal operating positionfor the reagent probe transport systems R1, R2 and R3 which are carriedby the plate 441. In this normal operating position, the reagentaspirating and dispensing probes for each of the systems R1, R2 and R3move forwardly and rearwardly between a rearward "home" position inwhich the probe is above a corresponding reagent dispense point and aforward aspirating position in which the probe is above a correspondingopening in the cover 327 of the reagent transport system. The plate 441is moved to the rearward position between test runs in order to positionthe guard which extends in front of the reagent probe transport systemsin back to the cover 327 of the reagent trays to enable the cover to beremoved for replacement of the reagent containers. The forward andrearward positions of the plate 441 arm determined by the sensors 448and 450 and a tab 431 which ends upwardly from the bracket 488. When theplate 441 reaches its rearward position, the tab 431 passes between theelements of the sensor 450 to interrupt a right beam and provide asignal to the CPU that the plate 441 is properly positioned at therearward position of the plate. When the plate 441 is in its forwardposition, the tab 431 is located between the elements of the sensor 449so that the beam which passes from one element to the other isinterrupted to provide an electrical signal to the CPU that the plate isproperly positioned in its forward position.

Referring particularly to FIGS. 63 and 64, the carriage 464 of thereagent probe transport system R1 includes a rear vertical wall 508which has a horizontal bore 511, a top wall 509, which has a verticalbore 514 and a bottom wall 510 which has a vertical bore 515. A bearing517 is located in the bore 515 and a bearing 521 is located in thevertical bore 514. A follower mounting guide 518 is fixed to the wall508 and has a cylindrical portion 516 which extends into the bore 511. Ahorizontal bore 513 extends through the mounting guide 518 and there isa pair of bearings 427 at each end of the bore 513. A lead screw 499 isfixed to the drive shaft 472 of the motor 468 by a coupling 500. Thelead screw 499 extends through a roll nut 501 in a bore 502 of a block503. The block 503 is pivotally mounted between a pair of parallel armsof a yoke 506 in the identical manner as the mounting of the block 493in the yoke 494 as shown in FIG. 67. The yoke 506 has a laterallyextending shaft 507 which is supported within the bearings 427 andextends through the bore 513 of the mounting follower guide 518. Sincethe roll nut 501 is fixed to the block 503, rotation of the lead screw499 upon the actuation of the motor 468, causes the block 503 to moveaxially along the lead screw 499. This causes the carriage 464 to moveforwardly or rearwardly relative to the support plate 441, depending onthe direction of rotation of the lead screw 499.

Referring also to FIG. 72, a probe holding arm 519 is mounted to afollower guide 505. The follower guide 505 has a horizontal bore 520which contains a roll nut 521 which is located between and in axialalignment with the bearings 425 and 517 in the upper and lower walls 509and 510, respectively, see FIG. 64. The lead screw follower 505 has atab 433 which is slidably mounted in a vertical groove 432 of a verticalpost 522, see FIGS. 64 and 70. The post 522 has a lower horizontalflange 512 which is located below the bottom wall 510. The flange 512has a bore 523 which is vertically aligned with the bore 515. The upperend of the post 522 is fixed to a gear segment 524 which has a bore 525.The gear segment 524 has gear teeth 526 which extend radially about thecenter of the bore 525. The gear segment 524 is located above the topwall 509 so that the bore 525 is in axial alignment with the bore 514.The teeth of the gear segment 524 are in driving engagement with theteeth 631 of a horizontal plate 629 which is fixed to the plate 444 asshown in FIG. 60. When the carriage 464 is in its rear position, theprobe holding arm 519 faces to the left as viewed in FIG. 60. As thecarriage 464 moves forwardly, the gear segment 524 rotates about thevertical axis of the lead screw 527. This causes the probe supportingarm 519 to rotate approximately 90° from the leftwardly facing positionas shown in FIGS. 60 and 62 to a forwardly facing position. Referring toFIG. 22, this causes the probe 535 to move along a curved path which isindicated by the dot and dash line 428. The line 428 intersects thevertical axes of the dispensing point 45, wash station 15 and theopenings 329 and 338 in the clear plastic cover 327 of the reagent trayas shown in FIG. 22.

A stepper motor 528 is fixed to a rearwardly extending horizontal flange529 of the carriage 464. The motor 528 has a downwardly extending driveshaft 530 which is fixed to a pulley 531. A vertical lead screw 527 isrotatably mounted within the bearings 425 and 517 and is drivinglyengaged with the roll nut 524 of the follower 505. The lead screw 527extends through the bores 523 and below the flange 512. The lower end ofthe lead screw 527 is fixed to a pulley 533, which is drivinglyconnected to the pulley 531 through a timing belt 532. The inner surfaceof the timing belt 532 has a plurality of teeth which engagecorresponding teeth on the pulleys 533 and 531 to provide a precisepredetermined degree of rotation of the pulley 533 for each driving stepof the stepper motor 528 (teeth not shown). When the stepper motor 528is actuated for rotating the lead screw 527 in one direction, the probeholding arm 519 is moved upwardly. When the lead screw 527 is rotated inthe opposite direction, the probe holding arm 519 is moved downwardlyrelative to the upper and lower walls 509 and 510 and the post 522.

An interrupt sensor 571 is located at the top of the groove 432. Whenthe probe holding arm 519 is moved to its upper position, a beam in thesensor 571 is interrupted to provide an electrical signal to the CPUthat the probe 535 is properly positioned in its upper position. Thesensor 571 is mounted on a PC board 537 which is attached to the post522, see FIG. 64. A connector 540 connects the PC board 537 to thejunction J15 of the PC board 537.

Referring to FIG. 72, a PC board 534 is fixed to the probe holding arm519. The arm 519 also supports a first reagent probe 535, see FIG. 62.Referring to FIG. 64, a bracket 538 is fixed to the upper wall 509 ofthe carriage 464 and has a plurality of upwardly extending tabs 536 forinteracting with interrupt sensors 451 and 449 on PC board 446. Thesensor 451 is a "home" sensor which provides a signal to the CPU whenthe rearmost tab 536 interrupts a beam between the two elements of thesensor when the carriage is in its "home" or rearward position. When thecarriage is in the "home" position, the probe 535 is directly over acuvette at the reagent dispense point 45. The tabs 536 also interactwith the interrupt sensor 449 to insure that the probe 535 is locatedprecisely at each of its forward positions. If the probe 535 is properlypositioned, at any of the forward positions, the beam of the sensor 449will be aligned with a space between two adjacent tabs or to the outsideof one of the tabs. If the probe is not properly positioned, the beamwill be interrupted by one of the tabs and a signal will be sent to theCPU to stop the machine.

The forward positions of the probe 535 include the wash station 15 andthe openings 328 and 338 of the outer cover 327 of the reagent tray 27.For each reagent pickup cycle, the motor 468 is actuated for apredetermined number of half steps to move the carriage 464 forwardlywith the probe 535 in the upper position from the home position untilthe probe 535 is above the wash station 15. The motor 528 is actuatedfor a predetermined number of half steps to lower the probe 535 into thewash station 18 for a wash cycle. The probe 535 is then raised byreversing the stepper motor 528 for a predetermined number of halfsteps. The motor 468 is actuated for a predetermined number of halfsteps to move the carriage 464 forwardly until the probe 535 is abovethe opening 328 or the opening 338 in the outer cover 327. If the testprotocol requires that the tracer or labeled reagent and the solid phasereagent are to be picked up by the probe 535, the probe is moved to eachof the openings 328 and 338 in succession. At each position 328 or 338,the probe 535 is lowered by the motor 528. The lower position of theprobe 535 is determined by a capacitance fluid sensing electronics asdescribed for the aspirating step for the sample probe 407. Afteraspiration of a volume of reagent, the probe 535 is raised to its upperposition, whereupon the motor 528 is actuated for a predetermined numberof half steps to move the carriage 464 so that the probe 535 is abovethe other reagent opening or moved rearwardly so that the probe 535 isabove the reagent disperse point 45. The reagent aspirating anddispensing probe is then lowered into a cuvette which is beneath thepoint 45. The volume of reagent is then dispensed into the samplesolution in the cuvette. The probe 535 is then raised to its upperposition and moved to the wash station 15 for a wash cycle which isdescribed in detail in following section of the description. Afterwashing of the probe, the probe is ready to begin another aspirating anddispensing cycle. The speed of the motor 564 is controlled by the CPU inaccordance with the operating program. The probe 535 is lowered to apoint just above the surface of the sample in the cuvette and thenraised at a predetermined rate while reagent is dispensed into thecuvette. The probe 535 is raised at a rate which maintains the tip ofthe probe just above the rising surface of fluid in the cuvette. Thisprovides maximum uniform mixing of the sample and reagent and minimizessplashing of fluids. This procedure also minimize the introduction ofair bubbles into the reaction mixture. This procedure is followed forthe reagent probe systems R2 and R3 which are described hereinafter. Aconnector 572 is connected to the PC board 534 of the arm 519 through aflexible lead 578 and is connected to the PC board 537. The metallicprobe 535 is electrically connected to the connector 572 and forms partof the capacitance level sensing system.

Referring more specifically to FIGS. 63, 65 and 69, the carriage 465 ofthe reagent probe system R2 includes a vertical forwardly facing wall541, a top horizontal wall 542 and a bottom horizontal wall 543. Thewall 541 has a horizontal bore 549 with a bearing 544 at each end of thebore. The top wall 542 has a bearing 557 which is located in a verticalbore 556. The bottom wall 543 has a bearing 558 which is located in avertical bore 559. The bores 556 and 559 are vertically aligned. Thewall 542 also has a vertical bore 545 which is vertically aligned with avertical bore 546 in the bottom wall 543. An anti pivot rod 547 islocated in the bores 546 and 545 and has an upper threaded end 548 whichis threaded into the carriage supporting slide bar 462. A lead screw 550is connected to the stepper motor 471 through a coupling 551 and extendsthrough a roll nut 552 in a block 553. The block 553 is mounted in ayoke 554 in the same manner as the mounting of the block 493 in the yoke494 as shown in FIG. 67. Since the roll nut 552 is fixed within theblock 553, rotation of the lead screw 550 upon actuation of the steppermotor 471 causes the block 553 to move along the longitudinal axis ofthe lead screw 550. The yoke 554 has a shaft 555 which is mounted withinthe bearings 554 and extends through the horizontal bore 549. As theblock moves forwardly and rearwardly along the longitudinal axis of thelead screw 550, it causes the entire carriage 465 to move forwardly andrearwardly relative to the support plate 441, depending on the directionof rotation of the lead screw 550 by the reversible stepper motor 471. Afollower guide 561 is located between the upper and lower walls 542 and543, respectively, and has a vertical bore 560 through which the antipivot rod 547 extends. Referring to FIG. 69, the follower guide 561 alsohas a vertical bore 574 which contains a roll nut 563. The follower 561is fixed to a probe carrying arm 562 which carries a reagent probe 576,see FIG. 62. A PC board 575 is connected to the arm 562, see FIG. 69. Avertical lead screw 573 is located within the roll nut 563 and isrotatably mounted Within the bearings 557 and 558. The bottom end of thelead screw 573 extends below the bottom wall 543 and is fixed to apulley 568. An electric reversible stepper motor 564 is fixed to a lowerand rearwardly extending horizontal bracket 565 of the carriage 465 andhas a downwardly extending drive shaft 566. A pulley 567 is fixed to theshaft 566 and is drivingly engaged. With the pulley 568 through a timingbelt 569. The interior surface of the timing belt 569 has teeth whichengage corresponding teeth on the pulleys 567 and 568, (teeth notshown). When the lead screw 573 is rotated in one direction by thestepper motor 564, the follower guide 561 moves upwardly relative to thesupport plate 441 along with the reagent probe 576. The reagent probe576 is moved downwardly with the follower guide 561 when the motor 564is reversed to rotate the lead screw 573 in the opposite direction. Anelectrical connector 570 extends from the stepper motor 564 and isconnected to the junction J13 on the PC board 446. A bracket 582 isfixed to the top wall 542 and has a plurality of upwardly extending tabs581 which interacts with the interrupter sensor 452 for insuring thatthe probe 576 is properly positioned at the several forward positions.If one of the tabs 581 interrupts a beam in the sensor 452 as any one ofthe forward positions of the probe 576, a signal is transmitted to theCPU that the probe is improperly positioned. A "home" tab 634 extendsupwardly from the carriage 465 and interacts with the interrupt sensor453. When the carriage 465 reaches its rearward "home" position, the tab634 interrupts the beam of the sensor 453 which transmits a signal tothe CPU that the carriage is properly positioned at the "home" positionin which the probe 576 is positioned over the reagent dispensing point46.

The stepper motors 471 and 564 are selectively controlled by the CPU tomove the carriage vertically and horizontally to position the probe 576in the same aspirating and dispensing sequence as described for theprobe 535 except that the probe 576 is moved in a straight forward toback line 426, see FIG. 22, which intersect the vertical axes of thereagent disposing point 46, the wash station 16, and the holes 339 and340 in the cover 327 of the reagent transport system 27. Depending onthe test protocol, the probe 576 will be moved forwardly to pick up oraspirate a labeled or tracer reagent at the opening 339 or a solid phasereagent at the opening 340. The test protocol nay also require that alabeled reagent and a solid phase reagent are to picked up by the probe576. The probe 576 is lowered by the motor 564 at each position 339 and340. The lower position of the probe 576 is determined by a capacitancefluid sensing electronics as described for the sample probe 407. Afteraspiration a volume of reagent, the probe 576 is moved to its upperposition, whereupon the motor 471 is actuated for a predetermined numberof half steps to move the probe above the other reagent opening orrearwardly so that the probe 576 is above the reagent dispense point 46.The probe is then lowered into a cuvette which is beneath the point 46.The aspirated-reagent is then dispensed into the sample solution in thecuvette. The probe 576 is then raised to its upper position and moved tothe wash station 16 for a wash cycle, whereupon it will be ready tobegin another aspirating and dispensing cycle.

Referring to FIGS. 22, 63, 66 and 71, the carriage 466 of the reagentprobe system R3 includes a rearwardly extending vertical wall 594, a tophorizontal wall 592 and a bottom horizontal wall 593. The vertical wall594 has a bore 595 which contains the cylindrical portion 580 of a guide608 which has a bore 579. A bearing 607 is located at each end of thebore 579. The top horizontal wall 592 has a bearing 590 which is locatedin a bore 591. The bottom wall 593 has a bearing 584 which is located ina bore 589. A lead screw 583 is rotatably mounted in the bearings 590and 584 and emends from the top wall 592 to the bottom wall 593. Thebottom of the lead screw 583 extends below the bottom wall 593 and isfixed to a pulley 600. A reversible stepper motor 596 is fixed to alower horizontally and rearwardly extending bracket 597. The motor 596has a downwardly extending drive shaft 598 which is fixed to a pulley599. The pulley 600 is drivingly connected to the pulley 599 through atiming belt 601. The inner surface of the belt 601 has teeth whichengage corresponding teeth on the drive pulleys 599 and 600 (teeth notshown). A reagent probe carrying arm 617 has a tab 627 which extendsinto a vertical slot in the rear side of the post 609 is fixed to a leadscrew follower 615 which has a roll nut 625 within a bore 616. The leadscrew 583 is drivingly engaged with the roll nut 625 for moving theprobe carrying arm 617 vertically up or down depending on the directionof rotation of the lead screw by the stepper motor 596. A vertical post609 is located between the upper wall 592 and the lower wall 593, andhas a lower rearwardly extending horizontal flange 610. The flange 610ends below the lower wall 593 and has a bore 611 which is verticallyaligned with the bore 589 so that the post is mounted on the bearing 584for rotation about the central longitudinal axis of the lead screw 583.The rear side of the post 609 has a vertical slot which is identical tothe slot 432 of the post 522. The reagent probe carrying arm 617 has atab 627 which extends horizontally into the vertical slot of the post609. This enables the post 609 to rotate with the gear segment 612 aboutthe longitudinal axis of the lead screw 583 for changing the angularposition of the third reagent probe 633 relative to the carriage 466. APC board 618 is fixed to the post 609 and has an interrupter sensor 624.An electrical connector 622 extends from the PC board 618 and isconnected to the junction J16 of the PC board 446. When the probecarrying arm 617 reaches its upper position, the tab 627 interrupts abeam on the sensor 624 which initiates a signal to the CPU whichindicates that the probe is properly positioned in its upper position.The back and forth motion of the carriage 466 is provided by the steppermotor 470 which has a drive shaft 474. The shaft 474 is fixed to a leadscrew 602 by a coupling 628. The lead screw 602 is engaged with a rollnut 603 in a block 604. The block 604 is mounted in a yoke 605 in thesame manner as block 493 which is mounted in the yoke 494 as shown inFIG. 67. The yoke 605 has a shaft 606 which is mounted in the bearing607 and extends through the bore 579 of the follower guide 608. Rotationof the lead screw 602 causes the block 604 to move along the centrallongitudinal axis of the lead screw. When the stepper motor 596 isrotated in one direction, the carriage 466 moves forwardly relative tothe plate 441. When the stepper motor 596 is reversed, the carriage 466is moved rearwardly relative to the plate 441. A bracket 620 is fixed tothe upper wall 592 of the carriage 466 and has a plurality of upwardlyextending tabs 621 which interact with the interrupt sensors 453 and454. The sensor 454 is a home sensor. When the carriage 466 is in itsrearward position so that the probe 633 is located above the reagentdispensing point 17, the rearmost tab 621 interrupts a beam in thesensor 454 which initiates a signal to the CPU that the probe is in its"home" position. The tabs 621 interrupt a beam in the sensor 453 whenthe probe 633 is improperly positioned in any one of its forwardaspirating or wash positions as described for the reagent probe systemsR1 and R2. A PC board 618 is fixed to the post 609 and has an electricalconnector 622 which is connected to the electrical junction J16 of thePC board 446. Referring to FIG. 71, a PC board 626 is fixed to the probesupporting arm 617 and is connected to the PC board 618 by an electricalconnector 619.

The upper end of the post 609 is fixed to a gear segment 612 which has abore 613. The gear segment 612 has gear teeth 614 which extend radiallyabout the center of the bore 613. The gear segment 612 is located abovethe top wall 592 so that the bore 613 is in axial alignment with thebore 613. The teeth of the gear segment 612 are in driving engagementwith the teeth 631 of a horizontal plate 630 as shown in FIG. 60. Whenthe carriage 466 is in its rear position, the probe holding arm 617faces to the right as viewed in FIG. 60. As the carriage 466 movesforwardly, the gear segment 612 rotates about the vertical axis of thelead screw 583. This causes the probe supporting arm to rotateapproximately 90° from the rightwardly facing position as shown in FIGS.60 and 62 to a forwardly facing position. This causes the probe 633 tomove along a curved path which is indicated by the dotted dot and dashline 429 as shown in FIG. 22. The line 429 intersects the vertical axesof the dispensing point 46, wash station 17, and the openings 341 and342 in the cover 327 of the reagent tray 27 as shown in FIG. 22.

Depending on the test protocol, the reagent aspirating and dispensingprobe 633 will be moved forwardly to pick up or aspirate a labeled ortracer reagent at the opening 341 or a solid phase reagent at theopening 342, see FIG. 22. Although the probe 633 is capable of pickingup labeled and solid phase reagent, the probe 633 is normally used forpicking up a single reagent The probe 633 is utilized for picking up areagent which compliments the single reagent which was picked up anddispensed into a cuvette by a preceding probe in accordance with aparticular test protocol. At each position 341 and 342, the probe 633 islowered by the motor 596. The lower position of the probe 633 isdetermined by a capacitance fluid sensing electronics as described forthe sample probe 407. After aspiration of a volume of reagent, the probe633 is moved to its upper position, whereupon the motor 470 is actuatedfor a predetermined number of half steps to move the probe above theother reagent opening or rearwardly so that the probe 633 is above thereagent dispense point 47. The probe is then lowered into a cuvettewhich is beneath the point 47. The aspirated reagent is then dispensedinto the sample solution in the cuvette. The probe 633 is then raised toits upper position and moved to the wash station 17 for a wash cycle,whereupon it will be ready to ben another aspirating and dispensingcycle.

The lower position of each reagent probe is determined by a capacitancefluid sensing system as described for the reagent probe systems R1 andR2.

In the preferred embodiment, the solid phase reagent and the labeledreagent are arranged in two separate concentric circles which maximizesthe number of reagent pairs that can be used with the analyzer. Thismean that each of the reagent probes must have two reagent aspiratingpositions in order to pick up either of the reagents. It is possible toplace the labeled reagent in the same type of container as the solidphase reagent and to place the container on the inner circle of holderswith the solid phase reagents. If a test protocol calls for bothreagents of a pair to be picked up by a probe, the probe would be raisedafter aspirating one of the reagents. This would allow the reagent trayto position the second reagent of the pair beneath the probe. The secondreagent would then be picked up by the probe.

Fluid Aspirating and Dispensing Apparatus

Referring to FIG. 73, the means for aspirating an dispensing fluidthrough the sample reagent probes includes the syringe bank 32 whichincludes a housing 650 and a plurality of stepper motors 655, 656, 657,and 658 which are mounted to the back of the housing 650. A plurality ofsyringes 651, 652, 653, and 654 are mounted to the front of the housingand are actuated by the stepper motors 655, 656, 657, and 658,respectively, the drive mechanism between each stepper motor and itsrespective syringe is a frictional rack and pinion drive which is shownand descrbd in U.S. Pat. No. 4,539,854 to Bradshaw et al. andincorporated herein by reference. Each syringe can be controlled toaspirate or dispense a small amount of fluid by controlling the signalsto the corresponding stepper motor from the CPU in accordance with themachine control program. The syringe 651 is operatively connected to thesample aspirating and dispensing probe 407 through a tube 659. Thesyringe 652 is operatively connected to the reagent aspirating andispensing probe 531 of the reagent probe system R1 through a tube 660.The syringe 653 is operatively connected to the reagent aspirating anddispensing probe 576 of the reagent probe system R2 by means of a tube661. The syringe 654 is operatively connected to the reagent aspiratingand dispensing probe 633 of the of the reagent probe system R3 by a tube662. Each tube which connects a reagent probe to its correspondingsyringe passes through a heated fluid bath 648. Each reagent probeaspirates a predetermined volume of reagent and after the probe has beenraised out of contact with the reagent solution the correspondingsyringe is operated for a predetermined draw of air which also draws theaspirated reagent into the fluid bath 648. The fluid bath 648 maintainsthe reagent at a predetermined operational temperature, preferably 37°C. A portion of the tube which is in the fluid bath is coiled so thatthe entire quantity of reagent solution is equilibrated to theoperational temperature before the reagent is dispensed into theappropriate cuvette. The air which has been drawn in behind the reagentis dispensed until the reagent reaches the tip of the probe prior todispensing of the reagent into the cuvette.

Referring to FIG. 75, wash stations 15, 16, 17, and 18 are shown mountedin front of the cuvette dispense and incubation section 39. Station 18comprises a tubular housing 666 which is mounted to the machineframework by a clamp 725. The housing 666 has a top opening 667, abottom outlet nipple 668 and a side port 669 which is located near thebottom opening 668. A tube 670 is connected to the nipple 668 and a tube671 is connected to the side port 669. The wash station 15 comprises atubular housing 663 which is mounted to the machine framework by a post688. The housing 663 has a top opening 673, a bottom outlet nipple 674and a side port 676 which is located near the bottom opening 674. A tube675 is connected to the nipple 674. A tube 677 is conned to the sideport 676. The wash station 16 comprise a tubular housing 678 which ismounted to the machine framework by a clamp 665. The housing 678 has atop opening 679, a bottom opening 680, and a side port 682 which islocated near the bottom outlet nipple 680. A tube 681 is connected tothe nipple 680 and a tube 683 is connected to the side port 682. Thewash station 17 comprises a tubular housing 684 which is fixed to a post691 which is fixed to the supporting base of the machine framework. Thehousing 684 has a top opening 685, a bottom outlet nipple 686, and aside port 687. A tube 690 is connected to the bottom opening 686 and atube 689 is connected to the side port 687.

Water supply to the wash stations from the reservoir 30 will bedescribed below.

The wash stations function to wash the various probes of the presentinvention between aspiration and dispense cycles. Deionized water isutilized as the wash solution in the preferred embodiment. Wash solutionis discarded in waste cover 31 after the wash cycle, as will bedescribed below.

Separation/Wash/Resuspend System

The reaction kinetics of the assays performed by the analyzer of thepresent invention are maximized by the elevated temperature and the veryefficient binding afforded by the large surface area of the paramagneticsolid-phase particles. Each assay sample undergoes the same totalincubation time of seven and one half minutes. When a cuvette reachesthe end of this total incubation time, it enters a section of theprocess track or incubation section where separation and washing isaccomplished. Powerful permanent magnets of neodymium-boron are mountedon the process track at this point, and the paramagnetic particles arerapidly pulled to the back wall of the cuvette. Liquid is aspirated fromthe cuvette by a vacuum probe which consistently seeks the bottom of thecuvette, the liquid being held in a waste reservoir for later disposal.Washing of the cuvette and particles is accomplished by forcefuldispensing of deionized water, followed by rapid magnetic separation andaspiration. One or two washes may be performed, based upon the specificassay, yielding non-specific binding of less than 0.1%. After completionof the wash cycle, the particles are resuspended in an acid containing0.5% hydrogen peroxide in a weak nitric acid, added from a fixed portabove the cuvette.

Referring to FIGS. 76-80, the aspirate resuspend area 28 includes ablock 694 which is mounted above the cuvettes and the aspirate resuspendarm at the downstream end of the cuvette dispense and incubation section39. A pair of spaced plumbing fixtures 695 and 700 are mounted in theblock 694. The fixture 695 has a bore 696 which extends completelythrough the block 694 to the cuvette and two tubes 697 and 698, whichcommunicate with the bore 696 and a nozzle 699 which extends through thefixture 695 in a fixed angular position. The nozzle 699 is connected toa tube 692 which is operatively connected to the reservoir 30 ofdeionized water. The nozzle 699 is positioned to direct a stream ofdeionized water against the front wall of the cuvette as shown in FIG.79. The fixture 700 has a bore 701 which extends completely through theblock 694 to the cuvettes and two tubes 702 and 703 which communicatewith the bore 701. An acid dispense fixture 704 is mounted to the block694 downsteam of the fixture 700. As shown in FIG. 80, a nozzle 706 ismounted in an angular fixed position in the fire 704 so that the end ofthe nozzle 706 is located just above the top opening of the cuvettewhich is positioned just beneath the fire 704. As shown in FIG. 80, thenozzle 706 is connected to a tube 707 which is operatively connected tothe acid reservoir 33, see FIG. 21B. The probe 706 is positioned at anangle to the vertical so that the stream of acid which is dispensed fromthe end of the nozzle is directed against the back wall of the cuvette40 for a purpose to be described.

Referring to FIG. 77, an aspirating unit which is generally indicated bythe reference numeral 708 is mounted on the fixed position behind theblock 694. The aspirating unit 708 comprises a fixed horizontalsupporting plate 709. A stepper motor 710 and a bracket 727 which aremounted on the plate 709. The bracket 727 has an upper horizontal flange714. A lead screw 717 is rotatably mounted in bearings 715 and 716 inthe flange 714 and the base 709, respectively. The lead screw 717extends through a roll nut 718 which is fixed within a bore 706 of afollower 719. The lower end of the lead screw 717 extends below the base709 and is fixed to a pulley 712. The drive shaft of the stepper motor710 extends below the base 709 and is fixed to a pulley 711. The pulley712 is driven from the pulley 711 through a timing belt 713 whichengages corresponding teeth on the pulleys 711 and 712, (teeth notshown). A forwardly extending arm 720 is fixed to the follower 719 andhas a pair of laterally extending arms 721 and 722. Referring also toFIG. 78, a probe 725 extends freely through the arm 721 and a housing723 which is fixed to the arm 721 and 725 has a protuberance 730 withinthe housing 723 which limits the upward movement of the probe relativeto the housing 73. The probe 725 is biased in the downward position by aspring 731. A probe 726 extends rely through the arm 722 and a housing724 which is identical to the housing 723 to limit the upward movementof the probe 726 relative to the arms 722 and the housing 724 and tobias the probe 726 downwardly. The probes 725 and 726 are verticallyaligned with the bore 696 and 701 respectively. Actuation of the motor710 the lead screw 717 to rotate about its vertical longitudinal axiswhich causes the follower 719 to move upwardly or downwardly dependingon the direction of rotation of the drive shaft of the stepper motor710. The vertical motion of the follower 719 causes the probes 725 and726 to move from an upper position in which the probes are above the topopenings of the cuvettes and a lower position in which the bottom tipsof the probes extend down to the bottom of the cuvettes. The arm 720 ismoved downwardly a distance which is slightly more than that which isrequired to enable the probes 725 and 726 to reach the bottom of thecuvettes. When the probes 725 and 726 strike the bottoms of theirrespective cuvettes, the additional slight movement of the arm 720causes the probes to move upwardly relative to the arms 721 and 722,respectively, against the bias of the springs 731. This guarantees thatthe bottom ends of the probes 725 and 726 will always be at the bottomof each cuvette for complete aspiration of the fluid in the cuvette. Thefollower 719 has a laterally extruding horizontal tab 744 which rides ina vertical slot 745 in the post 727. This prevents rotation of thefollower about the longitudinal axis of the lead screw 717. Aninterrupter sensor 746 is located at the top of the slot 745. When thefollower 719 reaches its upper position, the tab 744 interrupts a lightbeam between the two elements of the sensor 746 which initiates anelectrical signal to the CPU to indicate that the probes 725 and 726have reached their upper predetermined positions. At a designed time inthe machine operation sequence, the motor 710 is energized for apredetermined number of half steps to lower the probes 725 and 726 totheir lower positions.

Referring to FIG. 74, there is shown a cross-section of a heated tubeconfiguration which is generally indicated by the reference numeral 733.This configuration forms a portion of the tubing which connects eachreagent probe to its corresponding syringe that extends between theprobe and the heated fluid bath 648. The heated tube configuration 733comprises a teflon tube 734 through which the reagent flows, aninsulated heater wire 735 which is spirally wound around the tube 734and a thermistor 736. The tube 734, the heater wire 735 and thethermistor 736 are all enclosed within a shrink-wrap tube 737. Theheater wire 735 is a nickel--chromium wire which has a return lead 738outside of the shrink-wrap tube 737. The shrink-wrap tube 737 and thereturn lead 738 are, in turn, enclosed in a polyvinyl chloride tubing739. The function of the heated tube 733 is to maintain the temperatureof the reagent at 37° C. after it is transferred from the heated fluidbath, 648 to the reagent aspirating and dispensing probe. The CPUcontrols energization of the heater coil 735 in accordance withelectrical signals which are received from the thermistor 736 whichfunctions to maintain the temperature of the tube 734 at 37° C., plus orminus one degree. Although the heated fluid bath 648 is effective inheating the reagent to the desired predetermined temperature, i.e., 37°C., experience has shown that the temperature of the reagent drops belowthe predetermined set temperature as it passes back from the heatedfluid bath 648 to the reagent probe. The reason that this occurs is thatthe section of tubing between the reagent probe and the heated fluidbath is chilled by the reagent as it is aspirated from its container,particularly if the reagent is colder than room temperature, whichsometimes occurs at the beginning of the initial setup of a run of test.The prechilling of this section of the tube causes the tube to act as aheat-sink and absorb heat from the reagent when it passes back from theheated fluid bath 648. The heated tube configuration 733 maintains thetube at the set temperature and prevents this chilling effect. Thisinsures that the temperature of the reagent remains the same as it wasin the heated fluid bath 648. The entire structure of the heated tubeconfiguration 733 is flexible to compensate for the vertical movement ofthe reagent probe. The wall thickness of the teflon tube 734 is veryimportant for the satisfactory operation of the heated tubeconfiguration 733. The wall thickness of the teflon tube 734 is betweenand including 0.006 and 0.010 inches. If the wall thickness is below thelower value, the breakage frequency of the tube is consideredunacceptable. If the thickness is greater than 0.010 inches, theefficiency of heat transfer from the heater wire 735 to the reagentfluid as it passes through the tube 734, is significantly reduced,thereby making it difficult to maintain the reagent at the settemperature.

The tube 734 is made of a fluoroplastic material, specifically PTFE(polytetrafluorethylene). PTFE has exceptional resistance to chemicalsand heat and is used for coating and to impregnate porous structures.The relative stiffness or rigidity of PTFE renders it generallyunsuitable for fluid tubes. However, for the optimum thickness range ofthe tube 734, PTFE is sufficiently flexible and yet provides superiorheat transfer and chemical resistant qualities to the tube.

Referring also to FIGS. 34 and 35, the aspirate/resuspend area 28 alsoincludes three magnets 740, 741 and 742 which are located beneath thecuvette conveyor along the back wall of a channel 743 through which thecuvettes pass as they are carried by the drive belts 167 and 168. Eachof the magnets 740 and 741 is elongated and extend horizontally, seealso FIG. 21B. The magnet 741 extends from the end of the 740 on thedownstream side and is located at a slightly lower level than the magnet740 as shown in FIGS. 34 and 35. Each magnet 740 and 741 creates amagnetic field having a vertical north-south polarity. The magnet 742 islocated on the front wall of the channel 743 and extends downstream fromthe end of the magnet 741. The magnet 742 creates a magnetic fieldhaving a north-south polarity which is below the magnetic field of themagnet 741. As a cuvette enters the aspirate/resuspend area 28, theparamagnetic particles from the solid phase reagent are attracted towardthe magnet 740 and migrate to the back wall of the cuvette. As thecuvette continues to travel along the magnet 740, the paramagneticparticles begin to concentrate more towards the center of the magnet740. As the cuvette passes beneath the bore 696, the liquid in thecuvette is aspirated by the probe 725 and delivered to the waste fluidreservoir 31, while deionized water from the reservoir 30 is introducedinto the cuvette through the nozzle 699. The aspiration of the liquidfrom the cuvette effectively removes all of the unbound labeled reagentand unbound test sample from the sample reagent mixture. This processisolates the detectable product that is formed by the test reaction,i.e. the complex including the paramagnetic particles. The deionizedwater from the nozzle 699 is directed against the front wall of thecuvette to minimize any disturbance of the paramagnetic particlesagainst the back wall of the cuvette. As the cuvette advances from theposition beneath the bore 696 to the position beneath the bore 701, theparamagnetic particles continue to concentrate into a progressivelytightening mass or "pellet" against the back wall of the cuvette. Themagnet 741 is located in this area and since it is lower than the magnet740, the paramagnetic particles tend to congregate at a lower point inthe cuvette. This locates the concentrated mass of particles in an areawhich is below the level of the acid solution which is added in asubsequent step. When the cuvette stops at the point beneath the bore701, the probe 726 descends to the bottom of the cuvette and aspiratesthe wash solution of deionized water which is delivered to the fluidwaste reservoir 31. When the cuvette is next positioned beneath the bore705 of the fixture 704, the nozzle 706 dispenses a volume of an acidsolution such as hydrogen peroxide from the acid reservoir 33. Becauseof the angle of the probe 706, the acid is delivered against the backwall of the cuvette just above the concentration of paramagneticparticles. This effectively washes the particles away from the back walland resuspends them in the acid solution. As the cuvette moves away fromthe bore 705, it passes along the front magnetic 742 which helps to pullsome of the paramagnetic particles away from the rear part of thecuvette toward the front. This helps to distribute the particles evenlywithin the acid solution. Since the probes 725 and 726 are linked intothe same actuating mechanism, they are lowered into the bore 696 and701, respectively, simultaneously. While the probe 725 aspirates asample reagent solution from a cuvette beneath the bore 696, the probe726 aspirates a solution from a cuvette which is located beneath thebore 701. At the same time, the probe 706 dispenses a volume of acidsolution to a cuvette which is located downstream of the cuvette whichis located beneath the bore 701. The cuvette which is beneath the acidprobe 706 is then advanced toward the elevator mechanism to theluminometer which is described in the next section.

Luminometer System

The luminometer includes a rotary housing with six wells. A detectorincludes a photomultiplier tube (PMT) which is mounted in front of thehousing. A cuvette enters one of the wells in the housing from theentrance opening and is moved in increments to the exit opening. At thethird position from the entrance opening, the cuvette is aligned withthe PMT. This design effectively eliminates ambient light from themeasuring chamber prior to initiating the chemiluminescent reaction.With the cuvette positioned in front of the PMT, a base solution,containing dilute sodium hydroxide, is injected into the cuvette. Forone particular assay, for example, this causes the oxidation of anacridinium ester label and results in the emission of light photons of430 nm wavelength. This emission is a sharp spike within one second andhas a duration of 34 seconds. The intensity of the emission is measuredover a 5 second interval by the PMT, which operates in thephotoncounting mode. "Dark counts" are measured before the lightemission, and are subtracted automatically.

The luminometer system is shown in FIGS. 76 and 81-86 and comprises aluminometer assembly which is generally indicated by the referencenumeral 760 which is mounted on top of an elevator assembly which isgenerally indicated by the reference numeral 761. The elevator assembly761 comprises a housing 762 which has a vertical bore 763 which extendsfrom a chamber 764 at the end of the event conveyor to the luminometerassembly. Referring particularly to FIG. 83, the elevator assembly 761also includes a top plate 765 and a lower plate 766. A lead screw 767 isrotatably mounted in bearings 768 in the lower and upper plates 766 and765, respectively. A follower 769 is mounted on the lead screw 767 formovement along the central longitudinal axis of the lead screw upwardlyor downwardly depending upon the direction of rotation of the leadscrew. Plunger 771 is located below the chamber 764 and is fixedlyconnected to the follower 769 by a horizontal arm 770. A verticalanti-pivot rod 772 is fixed to the bottom plate 766 and the upper plate765 and extends freely through an aperture 780 in the arm 770. The lowerend of the lead screw 767 extends below the bottom plate 766 and isfixed to a sprocket 776. A stepper motor 773 is mounted to the lower endof the elevator assembly 761 and has a downwardly extending drive shaft774 which is fixed to a sprocket 775. The sprocket 776 is driven fromthe sprocket 775 through a drive chain 777, see FIG. 81. The motor 773is reversible. When the lead screw 767 is rotated in one direction thefollower 769 is moved from the lower position shown in full lines to theupper position shown in dotted lines in FIG. 83. This causes the plunger771 to move from the lower full line position to the upper dotted lineposition as shown in FIG. 83. When the lead screw 767 is rotated in theopposite direction, the follower 769 and the plunger 771 move downwardlyfrom the dotted line position to the full line position. The cuvettes 40are conveyed along the event conveyor at twenty second intervals. Everytwenty seconds a cuvette 40 is deposited into the chamber 764 from theevent conveyor while the plunger 771 is in the lower full line positionThe motor 773 is actuated for rotating the lead screw 767 so that theplunger 771 moves to the upper position carrying the cuvette 40 which isin the chamber 764 to the luminometer assembly 760. The follower 769 hasa horizontally extending tab which interacts with upper and lowerinterrupter sensors 758 and 759. When the follower is at the lowerposition shown in full lines in FIG. 83, the tab 778 interrupts a lightbeam between the two elements of the sensor 759 which initiates a signalto the CPU that the plunger 771 is properly positioned at the lowerposition. At a predetermined time in the overall machine sequence, acuvette 40 is delivered by the event conveyor to a point above theplunger 771 as shown in full lines in FIG. 83 and the motor 773 isenergized for a predetermined number of half steps to raise the plunger771 to the dotted line position which delivers the cuvette 40 to aposition within the luminometer assembly 760. When the follower 769reaches its upper position, the tab 778 interrupts a light beam betweenthe two elements of the sensor 758 which initiates a signal to the CPUthat the plunger 771 is property positioned at its upper position. Themotor 773 is then reversed for a predetermined number of half steps toreturn the plunger 771 to its lower position.

Referring particularly to FIGS. 83 and 84, the luminometer assembly 760comprises a bottom support plate 789 which is supported on the top plate765 of the elevator assembly. A luminometer housing 790 includes acylindrical vertical wall 788, a bottom wall 792 and a top wall 793. Thehousing 790 has a large circular chamber 791 which contains a carousel800. The luminometer housing 790 is supported on the bottom supportplate 789. The bottom plate 792 has a central uplifted portion 794 whichhas an aperture 795 which contains a bearing 796. The top wall 793 hasan aperture 799 which contain a bearing 798. A vertical shaft 797 isrotatably mounted in the bearings 796 and 798 and is fixed to a hub 787of the carousel 800. The upper end of the shaft 797 extends above thetop wall 793 and is fixed to a gear 801. A stepper motor 804 is mountedon the top wall 793 and has a downwardly descending drive shaft 803which is fixed to a gear 802. The gear 802 is in driving engagement withthe gear 801 for rotating the shaft 797 which causes the carousel 800 torotate about the central longitudinal axis of the shaft 797. An encoderwheel 805 is fixed to the top end of the shaft 797 above the gear 801. Aluminometer sensor board assembly 806 is fixed to the top wall 793. Theencoder wheel 805 has a plurality of spaced upwardly extending tabs 784which interacts with an interrupt sensor 783 which extends downwardlyfrom the PC board 806. In the embodiment shown in FIG. 84, there are sixtabs 784 which correspond to six external cavities or wells 814 in theouter wall of the carousel 800. The carousel 80 is indexed to a newposition every twenty seconds by the stepper motor 804 through the gears801 and 802. The stepper motor 804 is given an input signal from the CPUwhich causes the carousel 800 and the encoder wheel to rotated about theaxis of the shaft 797. The carousel continues to rotate until the edgeof one of the tabs 784 interrupts a light beam between the elements ofthe interrupt sensor 783. When this occurs, the motor 804 is deenergizedfor a predetermined time period, whereupon the motor will be energizedto move the carousel 800 to the new position. A side opening 807 islocated in the cylindrical vertical wall 788 and opens into a tunnel 810of a connector arm 809 which connects the luminometer housing 790 to aphotomultiplier tube 808. The bottom wall 792 has an entrance opening811 and an exit opening 812. The entrance opening 811 is verticallyaligned with the vertical bore 763 of the elevator assembly 761. Theexit opening 812 is vertically aligned with a waste receptacle 35 forthe cuvettes, see FIG. 21B. The six cavities 814 in the outer surface ofthe carousel 800 are sequentially vertically aligned with the openings811 and 812 as the carousel 800 is rotated about the axis of the shaft797. Each cavity 814 has an outer opening which is closed by thecylindrical wall 788 of the hub 780 and a bottom opening which is closedby the bottom wall 792. The upper wall of each cavity has a small accessopening 852 which leads to the cavity. The access openings 852 arecovered by the top wall 793 except when they are vertically aligned witha pair of holes 836 and 851 in the top wall 793 for a purpose to bedescribed. Referring to FIG. 86, as the carousel rotates about thecentral vertical axis of the shaft 797, relative to the housing 790,each cavity 814 is maintained light tight from light from the outsideexcept where the cavity is aligned with one of the openings 812 and 811.Each cuvette is delivered by the elevator 761 into a cavity 814 which isaligned with the opening 812. The carousel is rotated 60° every twentyseconds. The cuvette is carried in a circle about the axis of the shaft797 until it reaches the opening 811 and falls into the waste receptacle35. Every twenty seconds, a new cuvette is delivered into a cavity 814and a processed cuvette is dropped through the opening 811. The centraluplifted portion 794 forms a downwardly facing cavity 785. The upliftedportion 794 has an aperture 786 which faces the side opening 807. Areference LED (light emitting diode) 830 is mounted on a PC board 829.The PC board 829 is fixed to the bottom wall 792 so that the referenceLED 830 extends into the cavity 785. The LED 830 is periodicallyenergized to emit a beam of light and is positioned so that the beam oflight passes through the aperture 786 to the photomultiplier tube 808.The bottom opening of the cavity 785 is closed by a cover 831 so thatlight cannot enter the cavity from the outside. The amount of light fromthe LED is substantially greater than the light from a test flash and isbeyond the normal operating range of the photomultiplier tube 808. Alight filtering means, not shown, is positioned between the LED and thephotomultiplier tube 808 to alter or reduce the amount of light whichreaches the PMT from the LED.

Referring particularly to FIGS. 84 and 85, a wash/waste tower assembly816 is fixed to the tops of a plurality of vertical posts 815 which arein turn fixed to the bottom support plate 889. The assembly 816comprises a support plate 817 which is fixed to the posts 815, a steppermotor 818 and a post 819 which is fixed to the top of the plate 817. Thepost 819 has a laterally extending upper arm 820. A vertical lead screw823 is rotatably mounted in bearings 821 in the arm 820 and the plate817. A follower 824 is mounted on the lead screw 823 for movement alongthe central longitudinal axis of the lead screw. The lead screw isdrivingly engaged with a roll nut 813 which is mounted within thefollower 824. The stepper motor 818 has a downwardly extending driveshaft which is fixed to a pulley 826. The lower end of the lead screw823 extends below the plate 817 and is fixed to a pulley 825. The pulley825 is driven from the pulley 826 through a timing belt 827. The innersurface of the timer belt 827 has teeth which engage corresponding teethon the pulleys 825 and 826 (teeth not shown). Rotation of the steppermotor 818 in one direction causes the follower 824 to move upwardlyalong the lead screw 83 while rotation of the stepper motor in theopposite direction causes the follower 824 to move downwardly along thelead screw 823. A probe retainer arm 828 is fixed to the follower 824and extends forwardly and horizontally therefrom. The forward end of thearm 828 has a bore 833 which holds a probe assembly 832. The probeassembly 832 includes a housing 835 which is fixed to the arm 828 withthe bore 833 and an aspirating probe 834. The probe 834 is mounted inthe housing 835 for limited vertical movement and is biased in thedownward position in the same manner as the probes 725 and 726 asillustrated in FIG. 78. The upper end of the probe 834 is fixed to atube 836 which is operatively connected to the waste fluid reservoir 31.The follower 824 has a laterally extending arm 782 which rides in avertical groove 781 in the post 819 as the follower 824 moves verticallyrelative to the lead screw 823. The tab 782 prevents the follower 824from rotating about the central longitudinal axis of the lead screw. Aplumbing fixture 837 is mounted to the top wall 793 above the hole 836.The fixture 837 has a nozzle 838 which extends into the hole 836 and isconnected to a tube 839 which is operatively connect to the basesolution reservoir 34. A plumbing fixture 840 is fixed to the top wall793 just above the hole 851 and has a bore 841 which extends down to thehole 851. The probe 834 is vertically aligned with the bore 841 so thatwhen the probe is moved to its lower position, it enters the bore 841and extends through the hole 851 and through the access opening 852 ofone of the cavities 814 which is vertically aligned with the hole 851.The fixture 840 also has a pair of tubes 844 and 845 which areoperatively connected to the bore 841. The tube 844 is operativelyconnected to the deionized water reservoir 30 and the tube 845 isoperatively connected to the waste fluid reservoir 31. The upper end ofthe probe 834 is located in a housing 835 which is identical to thehousing 723 which is shown in FIG. 78. The probe 834 is programmed to belowered to the bottom of a cuvette which is located beneath the bore 841and slightly beyond. When the probe 834 reaches the bottom wall of thecuvette, it is forced upwardly relative to the housing 835 against thebias of the spring within the housing. This insures that the probe willalways reach the bottom of the cuvette for complete aspiration of fluidwithin the cuvette.

FIG. 86 is a diagrammatic representation of the bottom wall 792 and thephotomultiplier tube 808. The cuvette 40 is delivered by the elevator761 through the opening 812 in the bottom wall 792 to one of thecavities 814 which is aligned with the opening 812 and which isidentified in FIG. 86 as position 846. The cuvette is moved every twentyseconds in 60° increments in a circle about the axis of the shaft 797.The cuvette is moved from position 846 to position 847 and then toposition 848 in front of the opening 807. In this position, the nozzle838 delivers a predetermined volume of a basic solution 0.25N. NaOH tothe acid solution, eg. 0.1N. HNO₃ with 0.5% H₂ O₂, which is already inthe cuvette. This causes the generation of a chemiluminescent signal.The signal is detected over a five second interval by the PMT whichoperates in a photon-counting mode. A chemiluminescent signal or flashproduces a flash profile which is compared to a stored standard curve todetermine the analyte concentration in the sample. A masterdose-response curve is generated for each lot of reagents. Thisinformation is put into the analyzer by keyboard or bar code. Theinformation is calibrated by measuring two standards, whose values areused to adjust the stored master-curve. The recommended data ofreduction methods are selected from a spline fit, or four or fiveparameter logistic curve fits, and are preprogrammed for each assay. Thecuvette is next moved to position 849 which is beneath the bore 841. Theprobe 834 is lowered to the bore 841, the opening 851 and into thecuvette, which is beneath this position, through the access opening 852.All of the fluid contents in the cuvette are aspirated by the probe 834whereupon the probe 834 is raised to its upper position. The cuvette ismoved to position 850 and then moved toward position 853. When thecuvette reaches the opening 811, it fills through the opening and intothe cuvette waste receptacle 35.

Corrected counts are used to calculate analyte concentration in thesample using a stored master curve. At the time of manufacture of eachlot of reagents, a master dose-response curve is generated usingmultiple assay runs on multiple instruments. This lot-specificdose-response curve data is supplied with the reagents and input intothe CPU memory using an integral bar code-reading wand, or through thekeyboard. The stored master curve is recalibrated by assaying twocalibrators, whose values are predetermined and provided to thesoftware. Multi-analyte calibrators are provided for this purpose, andweekly recalibrations are recommended for most assays.

Reference LED Module for Chemiluminescence Assay

FIG. 87A-B, schematically illustrates the analyzer's LED module. Thereference LED utilizes optical feedback to provide a constant lightoutput which can be presented to the PMT.

The light output level may be set by adjusting an electronicallyadjustable potentiometer (EEPOT). This EEPOT is used to adjust the lightoutput for manufacturing and component variances. The EEPOT may be setwith a specific sequence of control signals, and is not designed forfield adjustment.

Advantageous features of the reference LED board are:

Compact packaging fits under the luminometer

Optical feedback yields constant 470 nm. calibration for thephotomultiplier tube signal

Compensated voltage reference for added stability

Electronically adjustable light output allows easy factory calibration

May be powered on/off from machine controller board

The power requirements of the preferred embodiments are:

    ______________________________________                                        for the Logic  +5.00 V +/- 5% (75 mA max.);                                   for the Analog +12.0 V +/- 10% (300 mA max.).                                 ______________________________________                                    

The unit is preferably configured as a 2.1" diameter two-sided board,with a ground plane on bottom side. The following connectors should beprovided:

a 5 pin pigtail connector to mate with the machine controller and powersource,

connection to luminometer home sensor board, and

a 4 pin header to facilitate programming of the EEPOT.

The Power Connector pigtail, J1, shown as in FIG. 87A-8 has thefollowing pin assignments:

    ______________________________________                                        Pin      Name                                                                 ______________________________________                                        1        LEDCTL (from machine controller, 0 = off, 1 = on)                    2        SB3 (from machine controller, not used)                              3        +5 V                                                                 4        +12 V                                                                5        GND                                                                  ______________________________________                                    

The EEPOT header Connector, J2 shown as in FIG. 87, has the followingpin assignments:

    ______________________________________                                        Pin      Name                                                                 ______________________________________                                        1        /INC        EEPOT wiper increment line                               2        UP/DOWN\                                                                        EEPOT direction select line                              3        /CS         EEPOT chip select                                        4        GND                                                                  ______________________________________                                    

The preferred embodiment of the reference LED circuitry is furtherdetailed in FIG. 87A-B. Because stray light from the LED could affectthe photomultiplier tube reading during sample analysis, the referenceLED can be turned off via a control line on the luminometer machinecontroller board. Q₁ and R₁ form the power control logic. (A in FIG.87A-B) Bringing LED CTL low (0 volts) turns off all op-amps and the LED;retuning LED CTL high turns the LED power on.

The closed loop that drives the LED uses a voltage as a command input(see FIG. 88). VR1, U1, U3A and R2, R3, and R7 comprise an adjustablevoltage reference. (B in FIG. 87A-B) VR1 provides atemperature-compensated zener reference of 6.9V +/-5%. The heater to VR1is on at all times to allow faster responses after instrument warm-up.R3, the EEPOT wiper resistance (10K), and R7 form a voltage divider.With the nominal values of these components, the EEPOT wiper has avoltage range of 0.5-2.5V. Op-amp U3A buffers the reference voltage toprovide a low-impedance source for the control loop.

An optical feedback loop is used to control the LED's light output CR1(blue LED, 470 nm wavelength) is a diffused bezel LED mounted in ahousing such that its light is incident upon the surface of CR2, abluesensitive photodiode. CR2 faces CR1 and is preferably positioned at45° off CR1's optical axis. The positioning of CR1 and CR2 is controlledby the LED mounting block. (Alternately a beam splitter may be providedto bring a portion of the LED output to CR₂). CR₂ is used in currentmode (virtual short circuit across its terminals) to eliminate darknoise in the reference.

Q2 and R6 are used to drive current through the LED; this current islimited to 50 mA by the values of the circuit components and the uppervoltage rail of U2. U2 alone cannot drive the LED at 50 mA.

FET-input op-amp U2 can tolerate inputs down to ground and can swing itsoutput from ground to about 3 volts off the positive rail. This groundoutput capability is important for operating the LED at low lightlevels. The FET-input capability was chosen to minimize effects of inputcurrent (Iin<30 pA) on the summing junction.

U2 works to maintain 0 volts between its input pins. This will force thevoltage across the series combination of R5 and R8 to be virtually equalto the reference voltage applied by U3A. The reference voltage acrossR5+R8 yields a reference current of 2.5-12.5 nA. In steady state, CR2'scurrent will equal the reference current; if CR2's current is constant,the light from CR1 causing that current is also constant.

In the event that the light output from CR1 fluctuates, the circuit'snegative feedback will correct the error. For example, if CR1 outputstoo much light, CR2's current will increase. This increase in currentwill flow through R4 and will drive Q2's base voltage down, causing theCR1's current to decrease. Similarly, too little light from CR1 causesU2 to output a higher voltage, yielding more current through CR1 andmore light output.

The response time of the circuit is limited by the combination of C5 andR4. C5 functions as an integrator to prevent any instantaneousfluctuation of the output, in effect averaging the error signal. R4 andC5 filter off any high frequency noise that would be superimposed on thelight output of CR1.

Because the current flowing through the reference resistors R5 and R8 ison the order of 10 nA, board leakage currents caused by flux and oilscan be have a detrimental effect. To prevent leakage currents fromdisturbing the circuit, the summing junction of the opamp should begiven special consideration. A teflon solder post C is provided to tieR5, CR2's anode, Us's summing input (pin 2), and C5 together. Anotherteflon post D is provided to join R5 and R8. Also, C5 should be a highinsulation resistance (>30000 Megohm) capacitor to minimize shuntleakage through the feedback path around U2. A third, non-insulated,solder post is used to provide a connection point for CR2's cathode.Finally, the entire assembly is cleaned very thoroughly and thenhermetically sealed to prevent deposits from forming.

In experimental testing, the circuit has shown that a short interval isnecessary to allow the circuit voltages and currents to stabilize. Aone-minute interval should be allowed between energization andobservation to ensure that the light output will be stable.

Test requirements:

In addition to the short circuit and open circuit tests performed by thein circuit tester, the following additional tests must be performed.

A. Power logic

With +12V and +5V applied to J1 pins 4 and 3 respectively, drive J1 pin1 to ground. Verify that no current flows through R6 and that thevoltage at U3 pin 1 is at ground potential. Now apply +12V to J1 pin 1.Verify that the voltage at pin U3 pin 1 is between 0.4 and 2.8 V.

B. EEPOT Logic

If the EEPOT'S non-volatile memory has a limited number of write cycles,varying this pot should only be done once during testing.

Bring the CS\pin to TTL (OV).

Next, apply pulses to the EEPOT'S INC pin and verify that the wipermoves in the direction of the U/D\pin. Vary the U/D\level and verifyEEPOT operation. Also, verify that the current flowing through R6changes with the value of the EEPOT setting. Timing information for theEEPOT'S control lines in the prefer embodiment is shown in FIG. 89.

C. Control loop

Because the summing junction carries such small currents, measurement atthis point is to be avoided. During the calibration of the LED and PMTmodule, the optical operation of the module will be verified.

Hydraulic and Pneumatic Controls

The hydraulic and pneumatic controls for the various subunits of theanalyzer are shown in FIGS. 90-93. All of the valves described hereinare electrically actuated via the CPU. Referring first to FIGS. 90, 91,93A and 93B, a pair of three way diverter valves V2 and V5 are connectedto a main water line 886 by a pair of flexible tubes 882 and 888,respectively. The main water line 886 is connected to the de-ionizedwater reservoir 30. A peristaltic pump 880 is operatively engaged withthe tube 882 for drawing water from the reservoir 30 to the valve V2. Aperistaltic pump 881 is operatively engaged with the tube 888 forpumping water from the reservoir 30 to the diverter valve V5. The valveV2 is connected to a three way diverter valve V1 by a tube 891 and to athree way diverter valve V3 by a tube 892. The diverter valve V5 isconnected to a three way diverter valve V4 by a tube 893 to a three waydiverter valve V6 by a tube 894. The valve V2 diverts water from thetube 882 to the valve V1, or the valve V3. The valve V2 is normallyclosed to the valve V1 and normally open to the valve V3. The valve V55diverts water from the tube 888 to the valve V4 or to the valve V6. Thevalve V5 is normally closed to the valve V6 and normally open to thevalve V4. The diverter valve V1 diverts water to the syringe 651 througha tube 890, or through the tube 671 to the housing 666 of the washstation 18, see FIG. 75. The valve V3 diverts water to the syringe 654through a tube 925, or to the housing 684 of the wash station 17 throughthe tube 689. The valve V5 diverts water from the tube 888 to the valveV4, or to the valve V6. The valve V4 diverts water to the syringe 652through a tube 895 or to the housing 672 of the wash station 15 throughthe tube 677. The valve V6 diverts water to the syringe 653 through atube 926, or to the housing 678 of the wash station 16 through the tube683. The valve V1 is normally closed to the tube 890 and normally opento the tube 671. The valve V3 is normally closed to the tube 925 andnormally open to the tube 689. The valve V4 is normally closed to thetube 895 and normally open to the line 677. The valve V6 is normallyclosed to the tube 926 and normally open to the tube 683. A check valve884 and a filter 883 is located in the tube 882. A check valve 902 and afilter 889 is located in the tube 888.

The waste fluid reservoir 31 is maintained at a sub-atmospheric pressureby a vacuum pump 896 which is connected to the waste fluid reservoir byan air line 897. A main air line 898 extends from the reservoir 31 andis connected to a manifold 899 by a tube 900. A plurality of valves V7,V8, V9, V10 and V11 are connected to the manifold 898 by tubes 910, 911,912, 913 and 908, respectively. A vacuum gauge 905 is also connected tothe manifold 898 by a tube 907. The valve V11 is a bleeder valve whichis opened and closed by a switch 906 which is, in turn, controlled bythe gauge 905. When the pressure in the manifold 899 exceeds apredetermined set pressure, as detected by the gauge 905, the switch 906is closed to open the bleeder valve V11 to release air and lower thepressure in the manifold 889 to the set pressure. When the set pressureis reached, the gauge 905 opens the switch 906 to close the valve V11.The valves V7, V8, V9 and V10 are on/off valves which are operativelyconnected to the wash stations 18, 15, 16, and 17, respectively. Thevalve V7 is connected to the bottom of the housing 666 of the washstation 18 by a tube 670. The valve V8 is connected to the bottom of thehousing 684 of the wash station 17 by a tube 690. The valve V9 isconnected to the bottom of the housing 672 of the wash station 15 by thetube 675. The valve V10 is connected to the bottom of the housing 678 ofthe wash station 16 by the tube 681.

A wash-dispense pump 903 is connected to the main water line 886 and tothe nozzle 699 by a tube 692. The pump 903 is a displacement pump whichis actuated by a motor 904. The pump 903 extends at an angle to thedrive shaft of the motor 904 and is connected to the drive shaft by auniversal coupling. The motor 904 is energized to rotate its drive shaftone complete revolution which produces a displacement cycle for thevalve 903. The amount of displacement is determined b the angle of thevalve relative to the drive shaft of the motor. When the motor 904 isactuated for a single displacement cycle, water is pumped from thereservoir 30 to the nozzle 699 of the fixture 695 for a wash cycle.

The main water line 886 is connected to a pair of on/off valves V16 andV18. The valve V16 is connected to a tube 909 which splits into thetubes 702 and 697, which are connected to the fixtures 700 and 695,respectively. The valve V18 is connected to the tube 844, which extendsfrom the fixture 840 at the luminometer assembly. The main vacuum line898 is connected to a manifold 901 and on/off valves V12, V13, V14, V15and V17 are connected to the manifold 901 by tubes 914, 915, 916, 917and 918, respectively. The valve V12 is connected to the tube 729 whichleads to the probe 725. The valve V13 is connected to the tube 728 whichleads to the probe 726. The valve V14 is connected to the tube 836 whichleads to the aspirating probe 834. The valve V15 is connected to a tube927 which splits into the previously described tubes 703 and 698 to thefixtures 700 and 695, respectively. The valve 17 is connected to thetube 845 which ends to the fixture 840. A low pressure switch 924 isconnected to the manifold 901 by a tube 919. When the pressure in themanifolds 901 and 899 falls below a predetermined minimum value, theswitch 924 sends a signal to the CPU to stop the machine.

A pump 920 is connected to the acid reservoir 33 by a tube 921 and tothe tube 707 which leads to the acid dispensing probe 706. A pump 922 isconnected to the base solution reservoir 34 by a tube 923 and to thetube 839 which extends to the base dispensing probe 838. Energization ofthe pump 920 dispenses a predetermined volume of acid from the reservoir33 through the nozzle 706. Energization of the pump 922 dispenses apredetermined volume of base solution through the nozzle 838. Referringparticularly to FIGS. 93A and 93B, a single cuvette 40 will be followedas it travels along the event conveyor and through the luminometer. Asample solution is obtained by positioning the sample aspirating anddispensing probe 407 above one of the openings 255 and 256 of the sampletransport system 26. The probe 407 is lowered into the sample containerand the syringe 651 is actuated with the valve V1 in the closed positionwith respect to the tube 90. This enables a volume of sample solution tobe aspirated by the probe 407. The probe 407 is then positioned over thesample dispense point 44 and lowered into a cuvette which is positionedbelow the point 44. The syringe 651 is then actuated to dispense theaspirated sample solution into the cuvette. Valves V1 and V2 areactuated to divert water to the syringe 651 for dispensing a smallamount of water into the cuvette to insure that all of the sample isdispensed. If the test protocol calls for the addition of a diluent orpretreatment solution, the housing 666 of the wash station 18 is filedwith water from the tube 671. The probe aspirates the diluent orpretreatment solution, moves to the wash station 18 and is dipped intothe water filled housing 666. The probe is then positioned over theselected test sample solution for lowering into the sample andaspirating a volume of sample. The probe is then moved to the sampledispense point 44 for dispensing the aspirated sample and diluentpretreatment solution into the cuvette. The cuvette then proceeds alongthe event conveyor toward the point 45. The sample probe 407 is thenmoved above the wash station 18 as water from the peristaltic pump 880is diverted from the valve V2 to the valve V1 which diverts the water tothe tube 890 which passes through the syringe 651 to the tube 659 and isdispensed through the probe 407 for cleaning the inside of the probe andthen diverted by the valve V1 through the tube 671 into the housing 666for washing the outside of the probe 407. The washing solution which isintroduced into the housing 666 by the probe 407 and the tube 671 isaspirated from the bottom of the housing through the tube 670 by openingof the valve V7. The initial dispensing of water through the probe 407fills the housing 666 which effectively cleans the outside of the probeas well. This water is aspirated from the bottom of the housing and thewater from the tube 671 provides a final cleaning to the outside of theprobe. The water is also aspirated from the bottom of the housing. Theaspirated fluid passes through the tube 910 into the manifold 899 andeventually to the wastewater reservoir 31 through the tubes 900 and 898.

After the cuvette 40 has been filled with sample at the sample dispenserpoint 44 it travels along the event conveyor to one of the reagentdispense points 45, 46, or 47, depending on tee protocol of the test.Each reagent aspirating and dispensing probe is capable of picking up oraspirating traces or labeled reagent from the outer ring and a solidphase reagent from the inner ring or only one of the reagents. Anycombination is possible. For example, for a particular cuvette, alabeled reagent may be picked up by the reagent probe system R1 whilethe solid phase reagent is picked up by the reagent probe system R2 orR3 when the cuvette is approximately positioned at either of thesesystems. On the other hand, the reagent probe system R1 can pick up asolid phase reagent while the labeled reagent is added by either thereagent probe systems R2 or R3. As a practical matter, the reagent probesystems R1 and R2 are used primarily for protocols which require theaspiration and dispensing of both reagent solutions by a single probe.Although the reagent probe system R3 is capable of aspirating bothreagents, less incubation time is available so that the system is usedprimarily for adding a reagent solution to a cuvette which contains asingle reagent that had been added by the reagent probe system R1 or R2.

If the test protocol calls for the aspiration of one or both reagents bythe reagent probe system R1, each reagent solution is aspirated by theactuation of the syringe 652 with the valve B4 closed with respect tothe tubes 895. The reagent or reagents are drawn into the coiled sectionof the tube 660 which lies in the heated fluid bath 648 by drawing airinto the probe 535 when the probe is out of contact with the reagentsolution. When the probe is positioned above the cuvette which containsthe corresponding sample to be tested, the syringe is actuated to firstdisplace the air which is in the tube 660 and thereafter to dispense thereagent solution into the cuvette. The probe 535 is then positioned overthe wash station 15 and then lowered into the wash station. The valve V4is actuated to divert water to the tube 895. The water flows through theprobe 535 for flooding the housing 672 and, simultaneously, washing theinside and outside of the probe 535. At the same time, the valve V9 isopened to aspirate the waste fluid from the bottom of the housing 672through the tube 675 which eventually finds its way to the waste fluidreservoir 31. The valve V4 is then returned to its normal state todivert water through the tube 677 into the housing 672 for a finalwashing of the outside of the probe 535. This valve V5 is in itsnormally open state with respect to the valve V4 for the washing cycleof the probe 535. If the test protocol calls for aspirating anddispensing of reagent by then reagent probe system R2, reagent isaspirated by the probe 576 by actuating the syringe 653 while the tube926 is closed with respect to the valve V6. The reagent is dispensedinto the cuvette which is located at the dispense point 46 by thesyringe 653 using the same procedures as for the reagent probe systemR1. The valve V5 is actuated to divert water to valve V6 and valve V6 isactuated to divert water through the tube 926 to the probe 576 when theprobe is positioned within the housing 678 of the wash station 16. Whenthe valve V6 is returned to its normally opened state to divert waterthrough the tube 683 for a final outside wash of the probe. The valveV10 is opened for aspirating all of the waste fluid from the housing 678through the tube 681.

If the test protocol calls for the introduction of a reagent by thereagent probe system R3, reagent is aspirated by the probe 653 byactuation of the syringe 654 with the valve V3 in its normally closedposition with resect to the tube 925. After dispensing of the reagentinto the cuvette by the probe 653 so the probe is positioned within thehousing 684 of the wash station 17 for a wash cycle With the valve V2 inits normally open position with resect to valve V3, the valve V3 isactuated to divert water through the tube 925 to the reagent probe 653for the initial washing step as described for the reagent probe systemsR1 and R2. Thereafter, the valve V3 is returned to its normal state sothat it is open with respect to the tube 689 for the final washing step.All of the waste fluid is aspirated from the bottom of the housing 684by opening of the valve V8.

The cuvette continues to be advanced along the event conveyor until itis positioned beneath the bore 696 of the fire 695. After the probe 725has been lowered, the probe 725 is lowered into the bore 696 so that itextends all the way to the bottom wall of the cuvette whereupon thevalve V12 is open for aspirating all of the liquid within the cuvette.The paramagnetic particles are drawn against the back wall of thecuvette by the magnets 740 and remain in the cuvette during aspirationof the liquid. The liquid includes unreacted labeled reagent andunreacted test sample. The pump 903 is actuated to dispense thedeionized water from the main line 886 through the nozzle 699 againstthe front wall of the cuvette. If the test protocol calls for a secondwash cycle, the deionized water from the first wash cycle is aspiratedthrough the probe 725 by again opening the valve V12. The pump 903 isactuated for a second time to introduce de-ionized water from the mainwater line 886 through the nozzle 699 for a second wash cycle. Theliquid from the second wash cycle or the first wash cycle if only onewash cycle is required, remains in the cuvette until the cuvette islocated beneath the port 701 of the fixture 700. When the probe 726 islowered through the bore 701 to the bottom of the cuvette, the valve V13is opened to aspirate all of the wash liquid from the cuvette. At thispoint all of the paramagnetic particles are held against the back wallof the cuvette by the magnets 741. When the cuvette arrives at a pointbeneath the acid dispense fire 704, the pump 920 is actuated to dispensea predetermined volume of acid from the acid reservoir 33 through thetube 707 and through the nozzle 706 against the back wall of the cuvettewhich dislodges all of the paramagnetic particles from the back wall andresuspends them into the acid solution.

After the addition of acid solution into the cuvette, the cuvette isadvanced along the event conveyor to the luminometer conveyor 761,whereupon the cuvette is raised to the luminometer 760. The cuvette isadvanced by the carousel 800 to the position 848 in line with theopening 807 which leads to the photomultiplier tube 808, see FIG. 86.With the cuvette in this position, the pump 922 is actuated to dispensea predetermined volume of base solution from the base reservoir 34through the nozzle 838. This produces a detection reaction "flash" whichis read by the photomultiplier tube 808 as described previously. Whenthe cuvette arrives at position 848 in the luminometer beneath the bore841, the probe 834 is lowered into the bore 841 to the bottom of thecuvette. The valve V14 is opened to aspirate the liquid in the cuvettethrough the probe 834 and through the tube 836 to the manifold 901. Theliquid is then drawn into the waste fluid reservoir 31. The valve 18 isthen opened to introduce water into the bore 841 while the valve V17 isopened. Continued aspiration of water through the probe 834 cleanses theinside of the probe while aspiration of water through the tube 845 helpsto cleanse the outside of the probe. When the cuvette is advanced to theopening 811 it falls through the opening into the waste receptacle 35.

All of the valves and pumps are controlled by the central processingunit in coordination with the operation of all of the machine subunitswhich are associated with the valves and pumps. All of the valves andother electrical components on the right side of the machine areconnected to a connector 928 by a ribbon cable (FIG. 92). The connector928 is operatively connect to the CPU. All of the valves and electricalcomponents on the left side of the machine are connected to a connector879 by a ribbon cable (FIGS. 90 and 91). The connector 879 isoperatively connected to the CPU.

Software Capabilities

The software system for the analyzer is capable of multitaskingoperation. At any time, the operator may access test results by sampleor by test, pending results by sample or by test, results history,calibration status, QC statistics, operating status, maintenanceschedule, or service history.

Test Definitions are custom programmable, including selection ofreporting units, number of decimal places in reported results, number ofreplicates, normal range, precision allowances, calibration interval,and automatic repeat with or without sample dilution.

Control Definitions are also programmable, including identity of controlselection of tests per control, and upper and lower limits per test,which will trigger flagging of out of range results. A plurality ofspecific test profiles may be defined and accessed. When a profile isrequested, all assays selected in that profile are automaticallyperformed.

Description of Flow Diagrams

FIGS. 94A and 95B constitute a single flow diagram and are connected bythe common symbol "PAGE 2". The diagram of FIGS. 94A and 94B is a timeline which illustrates the coordinated movements of the elements whichadvance the cuvettes from the supply hopper to the demotion point in theluminometer at the beginning of a test run. The diagram also depicts thecoordinated "home" or upper positioning of the probes and temperaturechecks. The designation "track" refers to the event conveyor and the"cuvette loader" refers to the mechanism for advancing the cuvettesalong the preheater section to the event conveyor.

FIGS. 95A, 95B and 95C constitute a single flow diagram. FIGS. 95A and95B are connected by their common symbol "PAGE". FIGS. 95B and 95C areconnected by their common symbol "PAGE 3" AND "PAGE 2A". The diagram ofFIGS. 95A, 95B and 95C is a time line which illustrated the coordinatedmovements of the mechanisms which advance the cuvettes and thecoordinated movements and functioning of the probes along the eventconveyor or "track".

FIGS. 96A, 96B and 96C constitute a single flow diagram. FIGS. 96A and96B are connected by their common symbol "PAGE 2". FIGS. 96B and 96C areconnected by their common symbol "PAGE 3". The diagram of FIGS. 96A,96B, and 96C is a time line diagram which depicts the coordinatedmovements of the elements which advance the cuvettes and thecoordination of the movements of the cuvettes with the dispensing ofsample and reagent into the cuvettes.

FIG. 97 is a time line which depicts the coordination of the movementsof the sample probe and the aspirating, dispensing and washing of thesample probe.

FIG. 98 is a time line diagram which depicts the coordinated movementsof the inner ring of the sample transport system and the sample probewhen a sample container or "cup" is added to the inner ring during a runof tests.

FIG. 99 is a time line diagram which depicts the movements of the probetransport system R1 in coordinating the functions of the probe for theR1 probe transport system.

FIG. 100 is a time line diagram which depicts the movements of the probetransport system R2 in coordination with the functions of the probe forthe R2 probe transport system.

FIG. 101 is a time line diagram which depicts the movements of the probetransport system R3 in coordination with the functions of the probe forthe R3 probe transport system.

FIG. 102 is a time line diagram which depicts the movements of theluminometer carousel and elevator in coordination with the functions ofthe luminometer.

Each subunit of the analyzer has its own routine which is determined bysoftware and microprocessor hardware. Each subunit routine is integratedby the CPU with interfacing hardware and software programs. Thecoordinated movements and functions of all the analyzer subunits aredetermined by software programming which functions through theelectronic hardware, reversible stepper motors, valves, pumps andsensors.

UTILITY OF THE INVENTION

A clinical laboratory instrument which is used to automate heterogeneousimmunoassay testing. The microprocessor-based instrument fully automateseach step of the assay.

It is obvious that minor changes may be made in the form andconstruction of the invention without departing from the material spiritthereof. It is not, however, desired to confine the invention to theexact form herein shown and described, but it is desired to include allsuch as properly come within the scope claimed.

EXAMPLES

The invention is further represented by the following examples whichdemodulate the operation of the analyzer. The examples are intended toillustrate the application of the analyzer for performing assays and notto limit the invention. It is to be understood that additional assays,including diagnostic and analytical, of various formats may beimplemented for use on the automated analyzer.

Example 1: Free Thyroxine (TT4)

A free thyroxine (FT4) assay has been developed for the above describedautomated analyzer. The FT4 assay is a competitive binding assay inwhich FT4 in a test sample competes with labeled T4 (tracer reagent) fora limited amount of T4 antiserum covalently coupled to the solid phase.In the preferred format of this assay acridinium ester is the label andparamagnetic particles serve as the solid phase. A test sample (25 uL.)acridinium ester labeled T4 (100 uL.) and anti - T4 paramagneticparticles (450 uL.) are dispensed by the analyzer into a cuvette andincubated for 7.5 minutes at 37° C. After incubation, magneticseparation and washes are performed as described prior to detection ofthe chemiluminescent signal. The amount of FT4 present in the testsample is determined by the level of the signed detected and isconverted to a dose by a two-point data reduction algorithm.

The test assay has a sensitivity of 0.107 ng/dL. (minimum detectabledose defined as the 95% confidence limit at 0 ng/dL.) with a range of0-13 ng/dL. The precision of the assay based on nine test runs overthree days is provided in Table 1. The correlation of the automated testassay with a manual test assay (Magic^(R) Lite Free T4, Ciba CorningDiagnostics, Corp.) provided a slope of 1.109, an intercept of 0.308 andcorrelation coefficient of 0.989 (N=131).

The specificity of the assay, i.e. % cross-reactivity, for variouscompounds is shown in Table 2.

                  TABLE 1                                                         ______________________________________                                        PRECISION                                                                     Based on 9 runs, 3 days                                                       Mean FT4                                                                      concentration,  Within   Total                                                ng, dL          run % CV % CV                                                 ______________________________________                                        0.62            4.5      5.1                                                  0.79            3.5      3.6                                                  1.05            3.5      7.9                                                  1.15            4.4      5.7                                                  1.39            3.5      4.4                                                  1.71            2.5      5.8                                                  6.42            4.7      5.9                                                  8.98            8.0      9.1                                                  ______________________________________                                    

                  TABLE 2                                                         ______________________________________                                        SPECIFICITY                                                                                          % Cross-                                               Compound               Reactivity                                             ______________________________________                                        L-triiodothyronine        3.9%                                                D-thyroxine            >64%                                                   D-triiodothyronine        3.6%                                                Diiodotyrosine          <0.002%                                               Monoiodotyrosine        <0.002%                                               3,5-diiodo-L-thyronine  <0.002%                                               Reverse triiodothyronine                                                                                3.1%                                                ______________________________________                                    

Example 2: Human Chorionic Gonadotropin (hCG)

A human chorionic gonadotropin (hCG) assay has been developed for theabove described automated analyzer. The hCG assay is a sandwich assaywhich utilizes an antibody-coated capture solid phase and a labeledantibody as a tracer reagent. In the preferred format of this assayacridinium ester is the label on a monoclonal antibody and polyclonalantibody coated paramagnetic particles serve as the capture solid phase.A test sample (50 uL.) and tracer reagent (100 uL.) are dispensed into acuvette by the analyzer and incubated for 5.0 minutes at 37° C. Thecapture solid phase reagent (450 uL.) is then added to the cuvettefollowed by an additional incubation of 2.5 minutes. After the secondincubation, magnetic separation and washes are performed as describedabove prior to detection of the chemiluminescent signal.

All data presented was generated based on a two-point calibration off afull standard master curve, consisting of ten standards. The standards,ranging from zero to 1000 mIU/mL., are calibrated against the WHO 1st751537 reference material.

The test assay has a sensitivity of less than 1 mIU/mL. (minimumdectable dose defined as the 95% confidence limit at 0 mIU/mL.) with arange of 0-1,000 mIU/mL. No hook effect seen at 40,000 mIU/mL. Theprecision of the assay based on five test runs over five weeks isprovided in Table 3. The specificity of the assay without cross reactantand with cross reactant is provided in Table 4. Interfering substancesadded to test samples according to NCCLS protocols were assayed withresults provided in Table 5. The correlation of the automated test assaywith a manual test assay with a manual test assay (Magic^(R) Lite hCG,Ciba Corning Diagnostics, Corp.) provided a slope of 1.08, an interceptof 1.03 and a correlation coefficient of 0.98 (N=172).

                  TABLE 3                                                         ______________________________________                                        PRECISION                                                                     Based on 5 weeks stored 2-point calibration, 5 runs                                   hCG      % CV of Dose                                                         Control, Within      Between                                          Study   mIU/mL   Run         Run    Total                                     ______________________________________                                        1       13.9     3.7         3.0    4.8                                               124.8    3.4         3.2    4.7                                               329.1    2.7         6.9    7.4                                       2       13.9     4.9         9.9    11.0                                              129.1    3.2         6.3    7.1                                               331.7    4.2         7.5    8.6                                       ______________________________________                                    

                  TABLE 4                                                         ______________________________________                                        SPECIFICITY                                                                               hCG result  hCG result                                            Cross       no cross    with cross                                            reactant    reactant,   reactant,                                                                              P value                                      (level tested)                                                                            mIU/mL      mIU/mL   (95% C.I)                                    ______________________________________                                        TSH         10.9        11.1     0.84                                         (2,000 uIU/mL)                                                                            207.0       214.9    0.26                                                     472.0       460.9    0.50                                                     832.8       812.0    0.68                                         FSH         13.1        13.4     0.35                                         (200 mIU/mL)                                                                              123.4       120.8    0.42                                                     431.5       427.6    0.16                                                     849.1       910.0    0.40                                         LH          4.5         4.5      0.85                                         (200 mIU/mL)                                                                              207.4       205.5    0.65                                                     459.1       480.2    0.10                                         ______________________________________                                    

                  TABLE 5                                                         ______________________________________                                        INTERFERING SUBSTANCES                                                        Patient samples were spiked with NCCLS recommended levels of                  various interfering substances. If P value > 0.05, the difference             in hCG dose is not statistically significant.                                         hCG        hCG      Spiked                                            Substance                                                                             Control,   Spiked,  vs.     P-Value (95%                              (mg/dL) mIU/mL     mIU/mL   Control C.I.)                                     ______________________________________                                        Conjugated                                                                            11.8       12.0        101% 0.54                                      Bilirubin                                                                             214.3      218.2    102     0.25                                      (20)    471.2      481.4    102     0.29                                      Unconjug.                                                                             2.7        2.9      106     0.34                                      Bilirubin                                                                             46.7       45.9      98     0.32                                      (20)    90.2       93.1     103     0.04                                              179.3      185.4    103     0.03                                              889.8      875.5     98     0.78                                      Lipid   2.9        3.1      107     0.54                                      (1,000) 22.0       23.1     105     0.12                                              48.3       50.5     105     0.04                                              94.3       98.7     105     0.00                                              191.7      189.8     99     0.57                                              571.1      934.4    107     0.31                                      Hemolysate                                                                            2.4        3.1      126     0.05                                      (500)   48.0       48.4     100     0.72                                              92.3       94.2     102     0.21                                              182.5      197.7    108     0.05                                              1,029.6    1,046.3  102     0.63                                      ______________________________________                                    

Example 3: Digoxin

A digoxin assay has been developed for the above described automatedanalyzer. The digoxin assay architecture is a hapten solid phase with alabeled antibody (tracer reagent). In the preferred format of thisassay, the tracer reagent is an acridinium ester labeled monoclonalantidigoxin antibody; and the solid phase is paramagnetic particles towhich digoxin-apoferritin has been immobilized. A test sample (150 uL.)and tracer reagent (50 uL.) are dispensed into a cuvette by the analyzerand incubated for 2.5 minutes at 37° C. The solid phase reagent (250uL.) is then added to the cuvette followed by an additional incubationof 5.0 minutes. After the second incubation, magnetic separation andwashes are performed as described above prior to detection of thechemiluminescent signal.

All data presented was generated based upon a two-point recalibrationoff an original master curve. The master curve was generated using eightstandards with values ranging from zero to 6 ng/mL digoxin.

The test assay has a sensitivity of less than 0.1 ng/mL (minimumdetectable dose defined as the 95% confidence limit at 0 ng/mL.) with arange of 0-5 ng/mL. The precision of the assay for patient samples andpatient pools is provided in Table 6. The specificity of the assay isprovided in Table 7. Interfering substances added to test samplesaccording to NCCLS protocols were assayed with results provided in Table8. The correlation of the automated test assay with a manual test assay(Magic^(R) Digoxin, Ciba Corning Diagnostics, Corp.) provided a slope of1.00, an intercept of 0.08 and a correlation coefficient of 0.97(N=130).

                  TABLE 6                                                         ______________________________________                                        PRECISION                                                                     ______________________________________                                        A. Patient samples run in replicates of two. 13 patient samples               were studied in each group.                                                          Mean digoxin                                                                           Within run                                                           concentration                                                                          % CV                                                          ______________________________________                                               0.52 ng/mL                                                                             6.5                                                                  0.81     4.7                                                                  1.05     4.7                                                                  1.22     4.9                                                                  1.37     5.6                                                                  1.49     5.2                                                                  1.86     4.2                                                                  2.68     2.3                                                           ______________________________________                                        B. Patient pools and control nm in replicates of 12 over 5 runs.              Digoxin                Within run                                                                             Total                                         concentration          % CV     % CV                                          ______________________________________                                        Controls: 0.79 ng/mL   7.0      7.9                                                     1.73         5.8      5.8                                                     2.81         4.8      5.0                                           Patient   0.62 ng/mL   6.7      8.0                                           Pools:    0.97         3.7      4.7                                                     1.15         5.1      5.5                                                     1.64         4.1      4.3                                                     2.05         4.3      4.6                                                     4.18         4.3      5.1                                           ______________________________________                                    

                  TABLE 7                                                         ______________________________________                                        SPECIFICITY                                                                   ______________________________________                                        Compound       % Cross-Reactivity                                             ______________________________________                                        Digitoxin      0.6%                                                           β-Methyldigoxin                                                                         109.4%                                                         Deslanoside    94.6%                                                          Digoxigenin    16.7%                                                          Lanatoside C   87.1%                                                          Ouabain        7.3%                                                           ______________________________________                                        Compound      Level Tested                                                                            Effect on Dose                                        ______________________________________                                        Cortisone     20 ug/mL  N.S.                                                  Estradiol     1 ug/mL   N.S.                                                  Progesterone  1 ug/mL   N.S.                                                  Testosterone  1 ug/mL   N.S.                                                  Prednisone    20 ug/mL  N.S.                                                  ______________________________________                                    

                  TABLE 8                                                         ______________________________________                                        INTERFERING SUBSTANCES                                                        Patient samples were spiked with NCCLS recommended levels                     of various interfering substances. If P value > 0.05, the difference          in digoxin dose is not statistically significant.                                        Digoxin Digoxin    Spiked                                                                              P-Value                                   Substance  Control,                                                                              Spiked,    vs.   (95%                                      (mg/dL)    ng/mL   ng/mL      Control                                                                             C.I.)                                     ______________________________________                                        Conjugated 0.003   0.008      --    0.36                                      Bilirubin  0.54    0.57       106%  0.20                                      (20)       2.23    2.21        99%  0.44                                      Unconjug.  0.004   0.000      --    0.30                                      Bilirubin  0.56    0.59       105%  0.06                                      (20)       2.25    2.22        99%  0.66                                      Lipid      0.010   0.012      --    0.89                                      (1,000)    0.52    0.58       112%  0.03                                                 2.06    2.04        99%  0.69                                      Hemolysate 0.0     0.0        --    1.00                                      (500)      0.52    0.53       102%  0.75                                                 2.09    2.10       101%  0.90                                      ______________________________________                                    

Example 4: Prostate Specific Antigen (PSA)

A prostate specific antigen (PSA) assay has been developed for the abovedescribed automated analyzer. The PSA assay utilizes an anti-PSAantibody solid phase and a labeled anti-PSA antibody as a tracerreagent. In the preferred format of this assay acridinium ester is thelabel on an affinity purified anti-PSA antibody and the solid phase isparamagnetic particles which is coated with anti-PSA monoclonalantibody. A test sample (100 uL.), tracer reagent (50 uL.) and solidphase reagent (250 uL.) are dispersed into a cuvette by the analyzer andincubated for 7.5 minutes at 37° C. After the incubation, magneticseparation and washes are performed as described above prior todetection of the chemiluminescent signal.

All data presented was generated based on a two-point calibration off astandard curve consisting of eight points.

The test assay has a sensitivity of 0.2 ng/mL. (minimum devable dosedefined as the 95% confidence limit at 0 ng/mL.) with a dynamic range of0-200 ng/mL. and a high dose hook capacity out to 40,000 ng/mL. Theprecision of the assay based on five separate runs on three instrumentsover a five day period for commercial controls and patient pools isprovided in Table 9. Interfringing substances, including endogenouscompounds and cheno therapeutic agents, added to test samples accordingto NCCLS protocols were assayed with results provided in Tables 10 and11. The correlation of the automated test assay with a manual test assay(Tandem R-R PSA, Hybritech) provided a slope of 1.01, an intercept of3.65 and a correlation coefficient of 0.97 (N=73).

                  TABLE 9                                                         ______________________________________                                        PRECISION                                                                     A. Analysis is based on 5 separate run on 3 instruments over a five           day period. Each run contained 12-14 repetitions.                             Two point calibration was used throughout                                     ______________________________________                                        Commercial                                                                              PSA            % CV                                                 Controls  Concentration, Within  % CV                                         (N = 70)  ng/mL          Run     Total                                        ______________________________________                                        A         2.76           8.7     11.15                                        B         7.71           6.74    7.36                                         C         17.37          5.94    6.91                                         ______________________________________                                        Patient   PSA            % CV                                                 Pools     Concentration, Within  % CV                                         (N = 60)  ng/mL          Run     Total                                        ______________________________________                                        1         15.79          4.49    6.46                                         2         25.91          5.73    7.64                                         3         48.78          5.54    8.65                                         4         93.66          5.81    8.07                                         ______________________________________                                    

                  TABLE 10                                                        ______________________________________                                        INTERFERING SUBSTANCES                                                        (ENDOGENOUS COMPOUNDS)                                                        Patient samples at various PSA levels were spiked with maximal levels         of endogenous interferents according to NCCCS protocols.                                 PSA     PSA        Spiked                                          Substance  Control,                                                                              Spiked,    vs.   Mean                                      (mg/dL)    ng/mL   ng/mL      Control                                                                             +/- SD                                    ______________________________________                                        Hemoglobin 7.08    7.32       103%   99                                       (500)      28.06   27.86       99%  +/- 4%                                               51.06   48.99       96%                                            Triglycerides                                                                            7.08    7.29       103%  102                                       (3000)     28.06   29.78      106%  +/- 5%                                               51.06   49.18       96%                                            Unconjug.  7.0     7.6        109%  103                                       Bilirubin  28.06   28.45      101%  +/- 6%                                    (20)       57.54   56.08       98%                                            Conjug.    7.08    7.57       107%  101                                       Bilirubin  28.06   29.44      105%  +/- 9%                                    (20)       51.06   46.57       91%                                            Total Protein                                                                            7.08    6.51        92%  90                                        (12 gm/dL) 28.06   25.38       90%  +/- 2%                                               57.54   50.98       89%                                            ______________________________________                                    

                  TABLE 11                                                        ______________________________________                                        INTERFERING SUBSTANCES                                                        (CHEMOTHERAPEUTIC AGENTS)                                                     Patient samples at various PSA levels were spiked with drugs                  commonly used in the treatment of cancer of the prostate (N = 5).                         PSA     PSA        Spiked                                         Substance   Control,                                                                              Spiked,    vs.   Mean                                     (ug/mL)     ng/mL   ng/mL      Control                                                                             +/- SD                                   ______________________________________                                        Cyclophosphamide                                                                          7.55    7.17        95%   98                                      (330)       28.06   27.52       97%  +/- 3%                                               49.34   49.8       101%                                           Doxorubicin 7.55    7.32        97%  100                                      (10)        28.06   28.22      101%  +/- 3%                                               49.34   50.11      102%                                           Megestrol   7.08    7.47       106%  101                                      Acetate     28.06   28.42      101%  +/- 5%                                   (79)        51.06   49.7       97%                                            Diethyl-    7.08    7.52       106%  101                                      Stilbesterol                                                                              28.06   28.10      100%  +/- 5%                                   (2.5)       57.54   55.57       97%                                           Methotrexate                                                                              7.08    7.16       101%  101                                      (13.2)      28.06   28.98      103%  +/- 3%                                               51.06   49.79       98%                                           ______________________________________                                         Prostatic acid phosphatase (PAP), >95% pure, showed less than 0.01% cross     reactivity                                                               

We claim:
 1. A container for reagents, comprising:a cylindrical bodyportion comprising inner and outer wall surfaces disposed substantiallyequidistant about a central axis and first and second open ends definedby said inner and outer wall surfaces, wherein a central axis mid-pointis defined between said first and second open ends; a floor memberattached to said first open end of said cylindrical body portion, saidfloor member having an inner surface; two co-planar agitating members,each attached to said inner wall surfaces of said cylindrical bodyportion and said floor member; a cylindrical neck portion attached tosaid second open end of said cylindrical body portion; and a skirtcomprising a radial portion, extending from at least a portion of saidouter wall surface of said cylindrical body portion in a directionperpendicular to said central axis, and an enclosing portion, attachedto said radial portion and having walls substantially equidistant tosaid central axis, said enclosing portion terminating on a side of saidfloor member opposite said first open end of said cylindrical bodyportion, wherein said two co-planar agitating members are disposed onopposite sides of said inner wall surface of said cylindrical bodyportion and extend, in a direction parallel to said central axis, fromsaid floor member to a point on said inner wall surface of saidcylindrical body portion between said mid-point and said second open endof said cylindrical body portion, and, in a direction perpendicular tosaid central axis, from said inner wall surface of said cylindrical bodyportion towards said central axis.
 2. The container of claim 1, whereineach of said first and second open ends of said cylindrical body portionlies in a respective plane, said respective planes being mutuallyparallel.
 3. The container of claim 2, wherein said central axis isorthogonal to each of said respective planes.
 4. The container of claim1, wherein said inner surface of said floor member is concave.
 5. Thecontainer of claim 1, wherein said cylindrical neck portion comprises athreaded outer wall surface.
 6. The container of claim 1, wherein saidradial portion of said skirt extends from a portion of said outer wallsurface of said cylindrical body portion adjacent said first open end ofsaid cylindrical body portion.
 7. The container of claim 1, wherein saidradial portion of said skirt extends from a portion of said outer wallsurface of said cylindrical body portion adjacent said second open endof said cylindrical body portion.
 8. The container of claim 1, whereinsaid two co-planar agitating members are each comprised of an inner edgesubstantially parallel to said central axis, said inner edges beingequidistant from said central axis.
 9. The container of claim 1, whereinsaid enclosing portion is substantially cylindrical.
 10. The containerof claim 1,wherein said enclosing portion further comprises first,second, third and fourth portions each having a first and second edgesubstantially parallel to said central axis, third edges incommunication with said radial portion, and fourth edges collectivelycomprising a terminating edge of said enclosing portion, wherein saidfirst, second and third portions are substantially planar and saidfourth portion is outwardly curved, and wherein said first edge of saidfirst portion and said second edge of said second portion are joined ata right angle, said first edge of said second portion and said secondedge of said third portion are joined at a right angle, said first edgeof said third portion and said second edge of said fourth portion arejoined, and said first edge of said fourth portion and said second edgeof said first portion are joined.